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Laboratory   Guide 


IN 


Experimental    Pharmacology 


DIRECTIONS  FOR  THE  COURSE  GIVEN  IN  THE 

University  of  Michigan 

BY 
CHARLES  W.  EDMUNDS,  A.  B.,  M.  D. 

j5^ 
INSTRUCTOR  IN  PHARMACOLOGY  IN  THE  UNIVERSITY  OF  MICHIGAN 

AND 

ARTHUR  R.  CUSHNY,  A.  M.,  M.  D. 

PROFESSOR   OF  PHARMACOLOGY  IN  UNIVERSITY   COLLEGE, 

LONDON;    AND     LATE    PROFESSOR     OF     MATERIA 

MEDICA    AND    THERAPEUTICS   IN   THE 

UNIVERSITY     OF    MICHIGAN 


GEORGE  WAHR 

PUBLISHER  AND  BOOKSELLER 

ANN  ARBOR,  MICH. 


COPYRIGHT   1905 

BY 
GEORGE  WAHR 


v, 


PREFACE. 

The  following  course  has  been  gradually  developed  in  the 
pharmacological  laboratory  of  the  University  of  Michigan, 
and  it  has  been  suggested  that  if  put  in  a  more  permanent 
form  than  hitherto  it  might  be  helpful  to  other  teachers  of 
this  subject. 

The  practical  course  has  hitherto  been  given  as  an  intro- 
duction to  the  didactic  lectures  and  it  has  been  found  advis- 
able to  limit  the  number  of  drugs  examined  to  those  possess- 
ing the  most  typical  action.  The  object  has  been  to  train  the 
student  entering  on  the  subject  in  the  method  of  work  and  to 
impress  on  him  that  the  study  of  drugs  is  to  be  approached 
in  the  same  objective  way  as  other  branches  of  medicine. 

The  order  in  which  the  experiments  are  arranged  is  a 
purely  arbitrary  one,  necessitated  by  the  conditions  of  the 
laboratory,  and  can  doubtless  be  modified  with  advantage  to 
suit  other  classes. 

A  few  days  are  devoted  to  purely  pharmaceutical  work, 
but  the  chief  weight  is  laid  on  the  experimental  part  of  the 
course.  The  few  hours  devoted  to  pharmacy,  however,  train 
the  student  in  observing  the  characters  of  the  much  larger 
number  of  drugs  met  in  the  lecture  course,  and  have  been 
found  to  be  of  considerable  value.  A  beginning  may  be 

3 


4  PREFACE. 

made  in  prescribing  as  occasion  offers,  but  this,  as  well  as  the 
therapeutic  applications  of  the  results  obtained  in  the  phar- 
macological experiments,  is  better  left  to  the  teacher  than 
expounded  in  the  laboratory  directions. 

ARTHUR  R.  CUSHNY. 
CHARTS  W.  EDMUNDS. 

Ann  Arbor,  March  i8th,  1905. 

We  wish  to  express  our  thanks  to  Mr.  Arthur  J.  Jones  for 
his  excellent  drawings  of  the  turtle,  the  dog's  head  and  the 
frog  with  its  heart  exposed. 


CONTENTS. 

PAGE. 

The  Pharmacopoeia   7 

General  directions  for  anaesthetics  and  operations n 

Chemistry   of    drugs : 24 

Physiology  of  frog's  nervous  system 44 

Soporifics    or    Hypnotics 51 

Percolation    59 

Nux  vomica 64 

Thujon    79 

Opium  80 

Curara 88 

Nicotine 92 

Veratrine    95 

Caffeine   96 

Cocaine  : 100 

Cinchona   103 

Emulsions   108 

Pills  112 

Digitalis   series    119 

Aconite   128 

Drugs  on  the  frog's  heart 132 

Drugs  on  the  rabbit's  heart  135 

Drugs  on  the  turtle's  heart 139 

Drugs  on  the  blood  pressure 147 

Drugs  on  the  dog's  heart 151 

Analysis  of  tracings 160 

Perfusion  of  the  kidney 167 

Diuresis   168 

Belladonna  series   179 

Drugs  on  the  sympathetic  system 187 

5 


6  CONTENTS. 

PAGE. 

Salivary   secretion    191 

The  sphygmomanometer  199 

Anaesthetics   204 

Pilocarpus  ....'.  208 

Suprarenal  extract 212 

Nitrite  series 216 

Intestinal   peristalsis    219 

Vasomotor  effects  in  the  intestine 223 

Vasomotor  effects  in  the  kidney 228 

Drugs  and  reagents  required 235 


The  Pharmacopoeia. 

When  a  physician  prescribes  a  certain  drug  from  time  to 
time  in  his  practice  it  is  of  greatest  importance  to  him  to 
know  that  his  patients  are  always  getting  the  same  prepara- 
tion, containing  the  same  active  principles  in  uniform 
strength.  These  requirements  could  not  be  carried  out  if 
there  were  not  some  standard  recognized  by  the  manufac- 
turers and  followed  by  them  in  the  preparation  of  the  various 
medicinal  remedies.  Most  countries  possess  such  a  standard 
in  their  " Pharmacopoeias,"  which  are  generally  published 
by  the  government.  In  this  country,  the  government  does 
not  issue  the  pharmacopoeia,  but  it  recognizes  its  au- 
thority. The  book  is  revised  every  ten  years  by  a  Committee 
of  Revision  composed  of  members  appointed  or  elected  by 
a  convention  of  various  medical  and  pharmaceutical  societies 
and  colleges. 

It  need  hardly  be  remarked  that  in  such  a  work  only  such 
remedies  are  included  as  have  become  well  established. 
These  are  designated  as  "official"  remedies,  while  drugs  not 
mentioned  are  "non-official." 

The  pharmacopoeia  gives,  first,  the  Latin  title  of  the  drug 
followed  by  the  English  name,  and  in  the  case  of  chemicals 
the  formula  and  molecular  weight  are  given.  Any  syno- 
nyms are  mentioned  and  wherever  it  is  necessary,  a  short, 
concise  definition  of  the  drug  is  added,  telling  its  nature  and 
its  source.  The  book  then  gives  tests  by  which  the  identity 
of  the  drug  may  be  recognized,  and  its  purity  and  strength 

7 


8  LABORATORY  GUIDE  IN 

tested  and  finally  mentions  in  what  ways  and  in  what  doses 
it  may  be  administered. 

Official  preparations  are  therefore  made  according  to  the 
directions  given  in  the  pharmacopoeias  and  they  should 
possess  uniform  strength  and  properties;  such  an'  ideal, 
however,  is  hard  to  attain,  as  some  of  the  crude  drugs,  even 
when  collected  according  to  pharmacopceial  directions,  dif- 
fer in  their  content  of  the  active  constituents. 


EXPERIMENTAL  PHARMACOLOGY. 


10  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  11 


General  Directions. 

Before  giving  the  specific  directions  for  the  experimental 
work  a  number  of  general  directions  are  given  for  the  com- 
moner operations  which  are  carried  out  on  animals  some  of 
which  are  usually  necessary  in  subsequent  manipulations. 

The  various  anaesthetics  employed  in  experimental  work 
are  also  described. 

Anaesthetics. 

No  animal  is  to  be  operated  on  unless  it  is  under  complete 
anaesthesia.  Ether  and  chloroform  may  be  used  for  this  pur- 
pose, but  have  the  disadvantage  of  requiring  constant  atten- 
tion. They  are  therefore  only  used  to  supplement  other 
anaesthetics  which  experience  has  shown  can  be  used  to  give 
full  anaesthesia  in  animals  without  the  necessity  of  their  being 
continually  administered.  For  this  purpose  the  following 
combinations  of  drugs  usually  give  very  good  results,  al- 
though in  rare  cases  chloroform  or  ether  may  have  to  be 
given  in  addition. 

Anesthetic  for  Rabbits. 

Paraldehyde,  1.7  cc.  per  Kg.  of  body  weight. 

While  your  assistant  holds  up  the  animal  by  all  four  legs 
and  head,  place  a  gag  in  the  mouth  and  pass  a  stomach  tube1 
through  the  opening  in  the  gag,  being  very  careful  not  to 
pass  it  into  the  lungs.  Draw  the  paraldehyde  into  a  pipette 


*A  stomach  tube  for  rabbits  and  cats  may  be  made  from  a  semi-elastic  catheter 
(No.  7  or  No.  8)  by  cutting  off  the  ivory  tip  and  fitting  in  its  place  a  short 
piece  of  rubber  tubing.  If  preferred,  the  soft  rubber  catheter  may  be  used. 


12  LABORATORY  GUIDE  IN 

and  place  the  point  of  the  pipette  in  the  opening  of  the 
stomach  tube  and  blow  the  drug  into  the  stomach,  and  then 
withdraw  the  tube. 

Anesthetic  for  Cats. 

Morphine  sulphate,  0.040  to  0.060  G.  Chloretone,  0.3  G. 
per  Kg.  of  body  weight. 

Place  the  animal  in  a  cat  box,1  draw  the  head  forward  and 
with  a  hypodermic  syringe  inject  the  morphine  solution 
under  the  skin  of  the  neck. 

Dissolve  the  chloretone  in  as  small  a  volume  of  alcohol  as 
possible  and  add  about  an  equal  volume  of  water.  Keep  the 
animal  in  the  box  and  introduce  this  solution  through  the 
stomach  tube  in  the  way  described  under  "Anaesthetic  for 
Rabbits." 

Anesthetic  for  a  Dog. 

Morphine  sulphate,  0.2  G.  to  0.25  G.  Chloretone,  3  G. 
to  40. 

Three  or  four  hours  before  the  animal  is  needed  inject  the 
morphine  solution  under  the  skin  by  means  of  a  hypodermic 
syringe. 

Just  before  operation,  dissolve  the  chloretone  in  alcohol 
and  add  about  an  equal  amount  of  water  and  give  it  to  the 
dog  by  means  of  a  stomach  tube2  passed  through  a  wooden 
gag. 


aA  cat  box  is  of  the  greatest  service  as  a  means  of  protection  to  the  operator 
whenever  this  animal  is  used.  It  consists  of  an  ordinary  wooden  box  about 
30  or  35  cm.  long,  18  cm.  wide  and  15  or  18  cm.  deep.  It  is  provided  with 
a  sliding  cover,  which  has  a  V-shaped  cut  in  the  middle  of  one  end.  In  the 
corresponding  end  of  the  box  a  second  V  cut  is  made.  When  the  lid  is  shut 
these  two  cuts  leave  enough  room  for  the  animal's  neck.  The  lid  is  kept  closed 
tightly  by  means  of  a  nail  passed  through  the  opposite  end  of  the  cover  into 
the  end  of  the  box. 

2The  average  stomach  tube,  such  as  is  used  in  clinical  medicine,  is  about  the 
right  size  for  a  dog.  It  should  be  about  75  cm.  long. 


EXPERIMENTAL   PHARMACOLOGY.  13 


14      .  LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  15 

Operations  on  Frogs. 

Pithing. 

Before  exposing  the  heart  of  a  frog  or  carrying  out  any 
other  operation  upon  the  animal  its  brain  is  destroyed 
(pithed)  in  the  following  manner:  Hold  the  frog  in  the 
left  hand  with  its  back  upward,  tip  the  head  forward  with  the 
left  index  finger,  and  feel  for  the  prominent  angle  made  at 
the  junction  of  the  skull  and  spinal  column.  Just  back  of  this 
angle  make  a  cut  with  small  scissors  through  the  skin  in 
the  median  line.  Now  push  the  sharpened  end  of  a  match 
forward  through  the  foramen  magnum  into  the  skull,  rotat- 
ing it  in  the  cranial  cavity  to  destroy  the  brain. 

Injecting  into  a  Lymph  Sac. 

In  the  intact  animal,  drugs  to  be  tested  are  usually  injected 
into  the  anterior  lymph  sac.  This  is  carried  out  in  the  fol- 
lowing manner :  Lay  the  animal  back  downwards  in  the  palm 
of  the  left  hand.  Hold  one  of  its  forelegs  firmly  between  the 
thumb  and  index  finger  and  the  other  foreleg  between  the 
middle  and  ring  fingers.  Draw  its  hind  legs  downward  and 
hold  them  against  the  palmar  surface  of  the  hand  by  means 
of  the  little  finger. 

Having  the  drug  in  the  glass  injecting  pipette,1  which  is 
held  in  the  right  hand,  force  the  animal's  mouth  open  with 
the  point.  Pass  the  pipette  into  the  mouth  avoiding  the 
tongue,  which  is  attached  anteriorly  and  direct  the  point  to- 


1To  make  a  glass  injecting  pipette  take  a  piece  of  glass  tubing  about  7  mm. 
in  diameter  and  about  20  or  30  cm.  long  and  heat  the  middle  point  in  the  Bun- 
sen  flame,  rotating  the  tube  constantly,  and  when  it  is  red  hot  remove  it  from 
the  flame  and  draw  it  out  to  a  capillary  size;  cut  it  in  the  middle  so  as  to  leave 
a  fine  tube  about  8  cm.  long.  Heat  the  large  end  of  the  pipette  to  smooth  it 
off  so  it  will  not  cut  the  tongue  or  lips. 

2-p 


16 


LABORATORY  GUIDE  IN 


ward  the  floor  of  the  mouth  which  with  a  little  pressure  it 
will  pierce,  entering  the  lymph  sac.  As  it  is  pushed  down  the 
sac  the  point  can  be  seen  beneath  the  skin  of  the  abdominal 
wall.  The  finger  is  now  removed  from  the  upper  end  of  the 
pipette  and  the  drug  allowed  to  flow  into  the  sac,  or  if  neces- 
sary blown  in. 


FIG.  No.   i. 

To  Expose  the  Heart. 

Pith  the  frog  and  tie  it  on  its  back  on  the  frog  board. 
With  a  pair  of  forceps  raise  the  skin  in  the  median  line  of 


EXPERIMENTAL  PHARMACOLOGY.  17 


18  LABORATORY   GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  19 

the  body  just  below  the  point  of  the  sternum  and  make  a  nick 
in  it  with  a  pair  of  scissors.  Starting,  at  this  point  cut  up 
and  outward  on  each  side  as  far  as  the  pectoral  girdle  and 
turn  this  V-shaped  piece  of  skin  upward  toward  the  head. 
Raise  the  sternum  by  placing  the  forceps  beneath  the  point 
and  with  scissors  cut  it  through  its  entire  length  avoiding  in- 
jury to  the  underlying  heart.  Tighten  the  strings  on  the 
fore  legs  so  as  to  pull  the  sides  of  the  sternum  well  apart. 
The  heart  is  now  seen  enclosed  in  the  pericardium,  which 
may  be  carefully  opened  and  cut  away. 

Directions  for  Operations  on  Mammals. 

The  following  are  the  most  common  operations  necessary 
in  experimental  pharmacology. 

To  Insert  a  Venous  Cannula. 
This  is  to  allow  of  the  intravenous  injection  of  drugs. 


FIG.   No.   2. 

Two  sizes  of  glass  venous  cannulas  (with  rubber  tubing 
attached)  are  shown  in  Fig.  No.  2.  The  larger  is  best  suited 
for  a  dog,  while  the  smaller  is  used  in  a  rabbit  or  cat.  Vari- 
ous sizes  should  be  provided. 

Operation.     When  the  anaesthesia  is  complete  make  an  in- 


20  LABORATORY   GUIDE   IN 

cision  through  the  skin  over  the  site  of  the  vessel  and  deepen 
the  cut  through  the  fascia  until  the  v.ein  is  exposed.  Avoid- 
ing injury  to  the  vessel  from  forceps  or  tearing,  clear  about 
two  inches  of  the  vein,  removing  all  the  fascia  surrounding 
it  so  as  to  leave  it  entirely  free.  Pass  two  threads  about  a 
foot  long  under  the  vein  and  place  "bull-dog"  forceps  on  it 
toward  the  cardiac  end  of  the  cleared  portion.  This  allows 
distention  of  the  vessel  by  the  blood.  Tie  one  of  the  threads 
tightly  around  the  upper  end  of  the  cleared  portion.  Hold 
the  end  of  this  ligature  in  the  left  hand  and  place  the  left 
index  finger  under  the  vessel,  thus  making  it  tense.  With 
sharp  scissors  make  a  cut  in  the  distended  vein  and  place  the 
tip  of  the  cannula  in  it,  and  tie  it  in  place  with  the  loose 
thread  previously  placed  around  the  vessel.  Fill  the  cannula 
and  vein  with  salt  solution  (0.8  %),  taking  great  care  that 
all  air  is  expelled. 

PRECAUTION.  When  drugs  are  injected  intravenously  it  is 
of  the  utmost  importance  to  see  that  no  air  is  allowed  to  enter 
the  veins.  A  small  bubble  of  air  is  almost  invariably  fatal 
to  rabbits.  Be  sure  the  cannula  and  syringe  have  all  the  air 
expelled. 

To  Insert  an  Arterial  Cannula.1 

This  is  to  allow  records  of  arterial  blood  pressure  to  be 
taken. 

Operation.  Expose  and  clear  the  artery,  usually  the 
carotid,  in  the  manner  described  above  for  the  vein.  Pass 
two  ligatures  under  the  vessel  and  tie  the  thread  which  is 
distal  to  the  heart.  Now  clamp  the  artery  with  the  "bull- 
dog" forceps,  placing  them  on  the  "heart  end"  of  the  cleared 


'•Arterial  cannulas  are  exactly  like  venous  cannulas.     Fig.  No.  2. 


EXPERIMENTAL   PHARMACOLOGY.  21 


22  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY. 


23 


portion.  Place  the  index  finger  under  the  vessel  as  before 
and  make  a  cut  in  it  and  tie  a  cannula  in  place  with  the  loose 
ligature.  This  cannula  is  to  be  rilled  with  sodium  sulphate 
solution  (page  147)  to  prevent  clotting  of  the  blood. 

To  Insert  a  Tracked  Cannula. 


FIG.  No.  3.      Glass  trachea!  cannula  with  rubber  tubing  attached. 

A  tracheal  cannula  is  inserted  to  allow  of  artificial  respira- 
tion being  carried  on  in  case  the  chest  is  to  be  opened  or  if 
the  anaesthesia  is  so  deep  that  the  respiratory  center  is 
seriously  depressed  or  paralyzed. 

A  convenient  form  of  tracheal  tube  is  shown  in  Fig.  3. 
They  can  be  made  of  various  sizes  of  glass  tubing  so  as  to  fit 
the  trachea  of  any  animal.  When  in  use  the  piece  of  rubber 
tubing  shown  in  the  cut  is  partially  constricted  to  give  the 
proper  degree  of  inflation  to  the  lungs. 

Operation.  Make  a  median  incision  in  the  neck  through 
the  skin  and  fascia  beginning  just  below  the  thyroid  cartil- 
age. Expose  the  trachea  by  separating  the  muscles  with  the 
fingers  or  blunt  hooks  and  dissect  away  all  structures  from 
around  it.  Pass  a  strong  thread  under  it  and  then  make  a 
transverse  cut  with  the  scalpel  between  two  rings  about  two- 
thirds  of  the  way  through.  Into  this  opening  pass  the 
tracheal  tube  and  tie  it  in  place  with  the  loose  ligature. 


24  LABORATORY  GUIDE  IN 


Chemistry  of  Drugs. 

Alkaloids. 

Among  the  most  important  of  the  organic  compounds  em- 
ployed in  medicine  is  the  group  of  vegetable  bases  known  as 
Alkaloids.  They  are  found  in  almost  all  parts  of  plants,  but 
in  greatest  abundance  in  the  seeds  and  roots.  They  consti- 
tute in  almost  all  cases  the  active  constituents  of  the  plants 
in  which  they  are  found  and  some  of  them  are  among  the 
most  powerful  poisons  known. 

The  alkaloids  contain  carbon,  hydrogen,  nitrogen,  and 
usually  oxygen,  but  in  a  few  instances  the  latter  is  not  pres- 
ent. 

In  their  chemical  properties  they  resemble  ammonia  very 
closely,  having  an  alkaline  reaction  and  in  uniting  with  acids 
without  the  elimination  of  hydrogen  to  form  salts.  These 
salts  differ  from  the  alkaloids  in  their  solubility,  and  upon 
this  fact  is  based  one  method  of  isolating  them  in  a  pure 
form  from  the  plants  in  which  they  are  found. 

One  of  the  most  familiar  alkaloids  is  quinine,  which  is  de- 
rived from  various  species  of  cinchona  bark,  and  it  may  be 
taken  as  a  type  of  the  group,  as  the  reactions  given  by  it  are 
characteristic  of  most  of  the  other  alkaloids. 

Taste  quinine  and  quinine  sulphate. 

I.  Test  the  solubility  of  a  few  crystals  of  quinine 
(alkaloid)  and  of  quinine  sulphate  (alkaloidal  salt)  in  10 
cc.  of  each  of  the  following: 


EXPERIMENTAL   PHARMACOLOGY.  25 


26 


LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY. 


27 


Solvent. 

Alkaloid  solubility. 

Alkaloidal  salt  solubility. 
Sts^fjjrf 

a 

Water. 



*~7/^ 

b 

Alcohol. 

*f~    *// 

se^-^Cci 

c 

Ether. 

^J^^c^y 

—  - 

d 

Chloroform. 

gu  <? 

— 

II.  Add  about  100  cc.  of  water  to  0.2  G.  quinine.     Shake 
the  flask  and  observe  only  a  small  part  goes  into  solution. 
Test  the  reaction  of  the  solution  with  litmus.     How  do  you 
explain  the  reaction  ? 

Add  i  cc.  dilute  sulphuric  acid  and  shake  until  complete 
solution  takes  place.  The  sulphuric  acid  formed  the  bisul- 
phate  of  the  alkaloid.  (Note  the  fluorescence  of  the  solution 
which  is  characteristic  of  quinine.)  Use  this  solution  in 
making  the  following  tests. 

III.  To  5  cc.  of  the  solution  add  slowly  potassium  carbon- 
ate solution  until   a   distinct   alkaline  reaction   is  obtained. 
What    happens?      Render    the    mixture    acid    with    dilute 
sulphuric  acid. 

Alkaloidal  salts  are  very  feeble  bases  and  are  thrown  out 
of  solution  by  the  addition  of  the  fixed  alkalies  and  their 
carbonates  which  unite  with  the  acid  of  the  alkaloidal  salt. 
The  precipitate  therefore  was  the  freed  alkaloid  which  is 
insoluble  in  water  and  which  on  the  addition  of  the  acid 
formed  the  sulphate  (or  bisulphate)  again,  which  passed 
into  solution. 

IV.  To  5  cc.  of  the  solution  add  potassium  hydroxide 
until  the  reaction  is  alkaline.    Add  distilled  water  until  the 
test  tube  is  nearly  full  and  then  shake  the  mixture  and  see  if 
the  precipitate  dissolves. 

The  precipitate  here,  as  in  No.  Ill,  is  the  free  alkaloid. 


28 


LABORATORY  GUIDE  IN 


The  alkaloids  are  spoken  of  as  being  insoluble  in  water,  but 
this  is  only  relatively  correct,  as  they  are  soluble  in  a  very 
large  excess;  in  the  case  of  quinine  in  1600  parts  of  cold 
water. 

In  the  first  parts  of  experiments  III  and  IV  it  was  seen  that 
an  alkaloidal  salt  is  thrown  out  of  solution  by  the  fixed  alka- 
lies and  their  carbonates.  For  this  reason  they  should  not 
be  prescribed  together  in  the  same  solution,  because  the  alka- 
loid would  fall  to  the  bottom  of  the  bottle  and  even  if  it  was 
shaken  each  time  before  a  dose  was  taken  the  chances  are 
that  the  last  dose  in  the  bottle  would  contain  a  much  larger 
amount  of  alkaloid  than  was  intended,  and  alarming  if  not 
fatal  symptoms  might  result.  The  same  objection  holds  in 
regard  to  prescribing  alkaloids  and  tannic  acids  together  in 
solution  (see  Exp.  Va). 

This  incompatibility  of  alkaloids  and  alkalies  may  be  over- 
come by  the  presence  of  a  sufficient  amount  of  alcohol. 

V.  Put  5  cc.  of  the  solution  into  each  of  5  test  tubes  and 
add  slowly  small  quantities  of  the  following  reagents;  note 
the  formation  of  any  precipitate  and  if  any  occurs  test  its 
solubility  by  adding  water  and  shaking  as  was  done  in  Exp. 
IV.  Tabulate  the  results. 


Reagent. 

Precipitate? 

Soluble  in  water? 

a 

Tannic  acid- 

-H^^ 

'     ~~   *y-A 

b- 

Picric  acid. 

.+—<&«<&£, 

r     ~*z&w 

c 

Iodine  in  potassium  iodide. 

-H^^; 

.«!  .. 

d 

Mercury-potassium  iodide. 

-^^r^tL 

*  * 

e 

Phosphotungstic  acid. 

*W^ 

•"r 

The  precipitate  in  (a)  was  the  tannate  of  the  alkaloid;  in 
(b)    the   picrate;    (c)    polyiodide    (usually);    (d)    double 


EXPERIMENTAL  PHARMACOLOGY.  29 


30 


LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  31 

iodide  of  the  alkaloid  and  mercury;  (e)  phosphotungstate  of 
the  alkaloid.  These  five  reagents,  with  a  few  others,  of  which 
the  most  important  are  gold  chloride  and  platinum  chloride, 
are  known  as  the  "alkaloidal  precipitants"  and  they  are 
largely  used  for  the  detection  of  the  different  members  of 
this  group. 

VI.  To   10  cc.   of  the  quinine  solution  add  potassium 
hydroxide  until  the  reaction  is  alkaline.     Now  add  5  or  10 
cc.  of  chloroform  and  placing  the  thumb  over  the  mouth  of 
the  test  tube  shake  it  vigorously  for  a  minute  or  two  or  until 
the  precipitate  disappears.     Set  it  aside  for  a  short  time  until 
the  chloroform  settles  in  the  bottom  of  the  tube  when  it  is  to 
be  drawn  off  from  the  water  by  means  of  a  glass  pipette  and 
put  into  a  clean  evaporating  dish  and  set  aside  to  allow  the 
chloroform  to  evaporate,  leaving  the  alkaloid. 

This  experiment  illustrates  one  of  the  methods  used  in 
separating  an  alkaloid  from  other  bodies  which  are  found 
present  with  it  in  the  crude  drug.  It  is  known  as  the  "  Shak- 
ing out  process"  and  will  be  more  fully  described  later.  (See 
nux  vomica  and  belladonna.) 

VII.  Boil  10  cc.  of  the  quinine  solution  for  a  few  min- 
utes with  a  cubic  centimeter  of  sulphuric  acid;  neutralize 
the  acid  with  potassium  hydrate  and  apply  Fehling's  test  for 
glucose.1     Is  there  any  reduction? 

Absence  of  reduction  shows  no  glucoside  is  present.  Con- 
trast next  section  (Glue  o  side s) . 


1Fehling's  test  for  glucose  is  carried  out  as  follows:  Mix  in  a  test 
tube  equal  quantities  (about  i  cc.)  of  two  solutions,  one,  a  solution  of  copper 
sulphate,  and  the  other,  a  solution  of  potassium  hydroxide  and  Rochelle  salts. 
The  resulting  mixture  is  a  clear,  dark  blue  solution,  which  is  to  be  boiled 
and  the  solution  to  be  tested  added  to  it.  If  glucose  is  present  a  red  precipi- 
tate of  cuprous  oxide  will  be  thrown  down  after  the  solution  has  stood  for  a 
short  time.  The  precipitate  may  be  clearly  seen  in  the  bottom  of  the  test  tube 
if  a  reducing  substance  is  present. 

3-p 


32  LABORATORY   GUIDE  IN 

Examine  specimens  of  other  alkaloids  and  their  salts  :— 

Strychnine,  from  the  nux  vomica  bean  (Strychnos  nux 
vomica). 

Morphine,  from  opium,  the  dried  juice  of  the  unripe  cap- 
sules of  the  poppy  (Papaver  somniferum). 

Caffeine,  from  the  berry  of  coffee  (Coffea  Arabica),  and 
from  tea  leaves  (Thea  Chinensis). 

Atropine,  from  deadly  nightshade   (Atropa  Belladonna). 

Berberine,1  from  golden  seal   (Hydrastis  Canadensis). 

Coniine,2  from  the  fruit  of  the  poison  hemlock  (Conium 
maculatum). 

Nicotine,2  from  tobacco  (Nicotiana  tabacum). 

Notice  that  the  names  of  alkaloids  end  in  "ine,"  in  Latin 
"ina,"  as  for  example,  strychnina,  morphina,  etc. 

Glucosides. 

Next  in  importance  to  the  alkaloids  among  the  vegetable 
poisons  rank  the  Glucosides.  These  are  substances  which 
break  up  when  acted  on  by  dilute  acids  or  ferments,  and 
yield  as  one  of  their  decomposition  products  a  sugar  which 
in  many  cases  is  glucose ;  in  fact,  glucosides  may  be  regarded 
as  salts  of  sugar.  The  other  products  differ  in  the  different 
glucosides,  in  some  cases  being  a  volatile  oil,  in  others  an 
alkaloid,  etc. 

In  the  following  experiments  amygdalin  will  be  employed. 
It  is  a  glucoside  which  is  found  in  bitter  almonds,  laurel 
leaves  and  in  the  bark  of  the  Virginian  prune  or  cherry. 

Make  50  cc.  of  a  2%  solution  of  amygdalin  in  water. 

I.     Apply  Fehling's  test  for  glucose  (see  page  31,  foot- 


1Berberine  is  one  of  the  few  colored  alkaloids. 

2Coniine  and  nicotine  are  the  two  most  important  liquid  alkaloids.     They  con- 
tain no  oxygen  and  are  volatile. 


EXPERIMENTAL  PHARMACOLOGY.  33 


34 


LABORATORY   GUIDE   IN 


/ 


EXPERIMENTAL  PHARMACOLOGY. 


35 


note)  to  5  cc.  of  the  amygdalin  solution.    Is  a  reducing  sub- 
stance present  ?  . 

II.  Heat^io  cc.  of  the  amygdalin  solution  with  I  cc.  of 
sulphuric  acid    (dilute)    on   a  water  bath   for   10  minutes. 
Neutralize  with  sodium  hydrate  and  apply  Fehling's  test. 
Do  you  get  any  reduction?     Compare  with  Exp.   I,  and 
explain. 

III.  Put  about  5  cc.  of  the  amygdalin  solution  into  each 
of  5  test  tubes  and  test  each  with  the  alkaloidal  precipitants. 


Reagent. 

Precipitate? 

a 

Tannic  acid. 



b 

Picric  acid. 

—  «- 

c 

Iodine  in  potassium  iodide. 



d 

Mercury-potassium  iodide. 



e 

Phosphotungstic  acid. 



IV.  Pulverize  six  bitter  almonds  in  a  dry  mortar  and  note 
any  odor  present.  Divide  the  powder  into  two  parts  placing 
one  portion  in  a  small  beaker  and  add  about  10  cc.  of  warm, 
not  hot,  water  to  it.  Boil  about  10  cc.  of  water  in  a 
test  tube  and  add  the  second  portion  of  the  powdered  almond 
to  it  and  boil  the  mixture  for  two  or  three  minutes.  Set 
them  aside  for  a  short  time  and  then  see  if  there  is  any  odor 
from  either  preparation.  Test  some  of  the  fluid  from  each 
with  Fehling's  solution  and  see  if  any  reduction  takes  place. 

In  the  bitter  almond,  in  addition  to  the  amygdalin,  is  also 
found  a  ferment,  emulsin.  Explain  therefore  the  presence 
of  a  reducing  agent  in  one  case  and  its  absence  in  the  other. 

The  odor  found  is  due  to  the  oil  of  bitter  almonds  formed 
by  the  decomposition  of  the  amygdalin  in  the  presence  of 


36  LABORATORY   GUIDE   IN 

water.  This  oil  consists  of  benzaldehyde  and  prussic  acid 
and  is  therefore  extremely  poisonous. 

What  reaction  is  characteristic  of  the  glucosides  ? 

Examine  specimens  of  other  glucosides : — 

Salicin,  found  in  the  willow  and  poplar. 

Digitalin,  found  in  the  fox  glove  (Digitalis  purpurea). 

Digitoxin,  found  in  the  fox  glove  (Digitalis  purpurea). 

The  names  of  glucosides  end  in  "in,"  in  Latin  "inum,"  as 
for  example,  salicinum. 

Volatile  Oils. 

The  group  of  volatile  oils  contains  a  large  number  of 
preparations  which  are  extensively  employed  in  medicine. 
They  are  also  known  as  the  Essential  or  Ethereal  oils.  They 
are  found  widely  distributed  in  nature  as  the  odor  of  the 
various  plants  and  flowers  is  due  to  them  or  to  their  oxidized 
products.  In  their  chemistry  they  differ  greatly  from  the 
group  of  fatty  or  fixed  oils  from  which  they  must  be  care- 
fully distinguished,  the  latter  being  non-volatile.  The  volatile 
oils  are  not  to  be  regarded  as  pure  substances,  like  the  alka- 
loids, but  rather  as  mixtures  of  varying  composition.  Most 
of  them  are  colorless  when  fresh  and  pure,  but  they  tend 
to  become  colored  after  long  exposure  to  light  and  air. 
In  odor  and  taste  they  differ  widely  from  one  another,  but 
in  both  cases  these  properties  are  generally  characteristic 
of  the  individual  oil.  The  majority  of  them  are  lighter  than 
water,  there  being  only  a  few  heavier. 

As  a  typical  member  of  the  group  the  oil  of  turpentine  is 
to  be  employed  in  the  following  experiments.  As  a  type  of 
the  fatty  or  fixed  oils  cotton  seed  oil  will  be  used  to  illustrate 
some  of  the  differences  between  the  two  series. 


EXPERIMENTAL  PHARMACOLOGY.  37 


38 


LABORATORY   GUIDE  IN 


*^^^(f,^)o     <^^f 


EXPERIMENTAL   PHARMACOLOGY. 


39 


I.     Solubility  of  volatile  oils. 

Into  each  of  five  test  tubes  put  I  cc.  of  the  oil  of  turpen- 
tine and  then  add  5  cc.  of  the  following  reagents.  Shake 
and  note  whether  the  oil  is  soluble. 


Reagent. 

Solubility? 

a 

Water. 



b 

Alcohol. 

•+~ 

c 

Ether. 

•+• 

d 

Chloroform. 

-/- 

e 

Cotton  seed  oil. 

+ 

II.     Solubility  of  fixed  oils. 

Into  each  of  5  test  tubes  place  i  cc.  of  cotton  seed  oil 
and  then  add  5  cc.  of  the  following  reagents.  Shake  and 
note  the  degree  of  solubility  as  in  I. 


Reagent. 

Solubility? 

a 

Water. 

— 

b 

Alcohol. 

f* 

c 

Ether. 

-/- 

-d 

Chloroform. 

f- 

e 

Oil  of  turpentine. 

^ 

What  differences  in  solubility  exist  between  the  two 
series?  ,•.  •  I 

The  two  groups  of  oils  resemble  one  another  in  leaving  a 
greasy  stain  when  dropped  on  paper,  but  that  from  the  fixed 
oil  is  permanent. 

III.  a.  Put  a  drop  of  oil  of  turpentine  on  glazed  paper. 
Note  the  stain.  Hold  the  paper  from  six  inches  to  a  foot 
above  the  flame  of  the  Bunsen  burner  for  a  few  minutes. 
Result  ? 


40  LABORATORY  GUIDE  IN 

b.  Repeat,  using  cotton  seed  oil,  being  careful  not  to  use 
enough  heat  to  char  the  paper.  How  does  it  differ  from  the 
previous  experiment? 

IV.  Decomposition  product. 

a.  Heat  5  drops  of  the  oil  of  turpentine  in  a  test  tube. 
Note  the  odor. 

b.  Heat  strongly  5  drops  of  cotton  seed  oil  in  a  dry  test 
tube.     Note  the  odor. 

In  the  latter  case  acrolein  is  formed  from  the  partial  de- 
composition of  the  fat  and  is  recognized  by  its  irritating 
and  disagreeable  odor  resembling  burnt  fat. 

V.  Add  i  cc.  oil  of  turpentine  to  5  cc.  water  and  shake 
the  mixture  well  for  a  minute  or  two.     Allow  the  oil  and 
water  to  separate,  and'  then  draw  of£  the  latter  with  a  pipette 
into  a  clean  test  tube.   Note  its  odor  and  taste,  which  are  due 
to  the  turpentine,  of  which  the  water  has  taken  up  a  small 
amount,  forming  a  preparation  recognized  in  the  Pharma- 
copoeia as  an  "Aqua."     These  waters  flavored  by  the  various 
volatile  oils  are  largely  used  to  give  a  pleasant  taste  and 
smell  to  medicines. 

The  "Syrupi"  are  solutions  of  sugar  and  water  to  which 
is  often  added  a  small  quantity  of  a  volatile  oil  as  in  the 
aquae. 

Spiritus,  spirits,  are  solutions  of  a  volatile  oil  in  alcohol 
(Exp.  I  b.)  ;  they  are  largely  used  in  medicine  as  flavoring 
agents. 

Examine  specimens  of  the  different  volatile  oils  named 
below  and  compare  them  with  the  oil  of  turpentine  as  re- 
gards odor,  taste,  color,  etc. 

Oleum  Menthae  Piperitaa  (oil  of  peppermint). 
Gaultherise  (wintergreen  oil). 
Lavandulse  Florum  (oil  of  lavender). 


EXPERIMENTAL   PHARMACOLOGY.  41 


< 


42  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  43 

Oleum  Eucalypti   (eucalyptus  oil). 
"       Caryophylli  (oil  of  cloves). 

Resins,  etc. 

Among  the  less  important  chemical  compounds  used  in 
medicine  are  the  Resins.  This  is  a  miscellaneous  class  char- 
acterized by  the  smooth,  shining  fracture.  As  a  class  they 
are  insoluble  in  water,  but  soluble  in  chloroform,  ether,  the 
volatile  oils  and  some  of  them  in  alcohol. 
Examine  the  Resin  of  jalap. 

Resin  of  copaiba. 

Oleoresins  are  solutions  of  resins  in  volatile  oils. 
Examine  Turpentine. 

Burgundy  pitch. 
Copaiba. 

Balsams  are  like  oleoresins,  but  contain  also  benzoic  and 
cinnamic  acids. 

Examine  Balsam  of  Peru. 
Balsam  of  Tolu. 

Gum  resins  are  mixtures  of  gums  and  resins,  containing 
usually  in  addition  some  volatile  oil. 
Examine  Asafoetida. 

Gums  are  amorphous  transparent  substances  belonging  to 
the  group  of  carbohydrates.     Some  dissolve  in  water  while 
others  only  swell  to  a  jelly.     They  are  insoluble  in  alcohol. 
Examine  Acacia. 

Tragacanth. 


44  LABORATORY   GUIDE   IN 


Physiology  of  the  Nervous  System  of  the  Frog. 

In  order  to  study  the  action  of  a  drug  upon  the  nervous 
system  of  a  frog  it  need  hardly  be  pointed  out  that  it  is  of 
prime  importance  to  understand  the  physiology  of  the  differ- 
ent parts  of  that  system.  To  illustrate,  a  certain  drug  will 
depress  the  cerebrum,  while  a  second  drug  will  stimulate  the 
spinal  cord  and  a  third  may  paralyze  the  peripheral  nerves ; 
now,  only  by  knowing  the  normal  functions  of  these  di- 
visions of  the  nervous  system  can  we  recognize  and  interpret 
the  symptoms  arising  from  an  increase  or  diminution  in  their 
activity,  and  thereby  locate  the  point  of  action  of  the  drug 
employed. 

First,  examine  the  normal  frog,  observing  its  spontaneous 
movements  and  the  method  of  leaping  and  what  degree  of 
irritation  is  necessary  to  make  it  move.  Note  also  its  co- 
ordination of  the  muscles  in  swimming;  its  recovery  of  bal- 
ance when  slowly  turned  on  a  sloping  board,  its  recovery 
when  placed  on  its  back,  its  reflexes  when  the  toe  is  pinched, 
the  croaking  when  its  back  and  sides  are  stroked,  the  rate  of 
respiration  and  heart  beat.  (The  latter  can  usually  be 
counted  when  the  animal  is  turned  on  its  back  and  placed  in 
a  good  light,  a  slight  shadowy  movement  of  the  skin  being 
seen  in  the  cardiac  region  with  each  pulsation  of  the  organ.) 

Having  studied  the  intact  animal,  separate  the  cerebral 
hemispheres  from  the  optic  lobes  by  thrusting  the  point  of 
a  scalpel  through  the  roof  of  the  skull  on  a  line  joining  the 
posterior  borders  of  the  eyelids,  gently  rocking  the  scalpel 
from  side  to  side  so  as  to  completely  sever  the  hemispheres 
from  their  inferior  connections.  As  will  be  seen  from  the 


EXPERIMENTAL   PHARMACOLOGY. 


45 


46 


LABORATORY   GUIDE  IN 


^ — ^Y      ^) 


EXPERIMENTAL   PHARMACOLOGY. 


47 


4     rog' 


accompanying  figure,  such  a  cut 
will  also  destroy  the  optic  thalami 
and  some  symptoms  will  result 
from  such  destruction,  but  for  all 
purposes  here,  the  differences  in 
results  obtained  are  not  important. 
After  allowing  the  animal  a  few 
minutes  to  recover  from  the  effects 
of  the  shock  of  the  operation 
(which  must  be  done  in  all  cases) 
compare  its  condition  with  that  of 
the  normal  animal. 

Make  full  notes  of  all  changes 
seen. 

Next  remove  the  optic  lobes 
from  the  same  animal  by  cutting 
as  before  with  the  point  of  a 


n,v        . 

Cerebral     hemispheres;    b.     Optic 

thalami;  c.  Optic  lobes,  d.  Cere-  scalpel  on  a  line  joining  the  cen- 

bellum.  e.  Medulla.     (Steiner) 

ters  of  the  tympanic  membranes. 

These  membranes  are  seen  on  the  sides  of  the  head  posterior 
to  the  eyes  as  two  flat  circular  surfaces  about  3  mm.  in 
diameter.  Wait  a  short  time  as  before  and  then  study  the 
effects  of  the  operation.  Hang  up  the  animal  by  means  of  a 
wire  hook  passed  through  the  lower  jaw  and  observe  the 
reflexes  on  pinching  the  toes  or  on  dipping  the  toe  in  very 
dilute  acid  (made  by  Adding  dilute  sulphuric  acid  to  water). 
Wash  off  the  acid  with  water. 

Using  the  same  frog  thrust  a  fine  wire  down  the  spinal 
canal  so  as  to  destroy  the  cord.  Observe  any  change  in  the 
reflexes.  What  is  the  condition  of  the  heart  and  respiration  ? 

Examine  the  activity  of  the  peripheral  nerves  in  the  follow- 
4-p 


48  LABORATORY   GUIDE,   IN 

ing  manner;  hold  the  frog  back  upwards  in  the  hand  and 
apply  the  electrodes  from  the  induction  coil  (tetanizing  cur- 
rent) to  the  lumbar  spine.  Observe  how  long  the  muscles 
will  stay  contracted  before  fatigue  sets  in. 

Dissect  out  the  sciatic  nerve  by  an  incision  along  the  pos- 
terior surface  of  the  thigh  and  stimulate  it  with  the  electric 
current  as  before. 

Lay  a  few  crystals  of  sodium  chloride  on  the  nerve  and 
moisten  them  with  a  drop  of  water.  Observe  any  change 
in  the  muscles.  Explain. 

Expose  the  gastrocnemius  muscle  and  stimulate  it  directly 
with  the  electric  current  and  also  with  salt  crystals. 

Describe  the  functions  of  the  different  parts  of  the  nervous 
system  in  the  frog  as  learned  from  the  symptoms  arising 
from  their  removal. 


EXPERIMENTAL   PHARMACOLOGY. 


49 


50  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY. 


51 


Soporifics  or  Hypnotics. 

Soporifics  or  Hypnotics  are  drugs  which  are  used  to  pro- 
duce sleep.  They  comprise  a  very  large  and  important 
group  of  substances  which  are  extensively  employed  in  medi- 
cine. In  their  action  they  are  closely  related  to  chloroform 
and  ether,  but  they  are  not  used  to  produce  anaesthesia  in 
man  owing  to  the  fact  that  it  is  impossible  to  regulate  suf- 
ficiently accurately  the  depth  of  the  anaesthesia  induced  by 
them. 

Chloral  (chloral  hydrate),  the  most  important  member  of 
the  'series,  was  introduced  into  medicine  by  Liebriech  in 
1869,  and  as  a  pure  hypnotic  drug  is  probably  the  most  re- 
liable in  use  today.  The  other  members  of  the  group  which 
are  of  greatest  importance  are  Sulphonal  and  Paraldehyde. 

Examine  each  drug  as  to  appearance,  odor  and  taste. 

i.    Examine  the  solubility  of  the  following  soporifics: 


Chloral. 

Sulphonal. 

Paraldehyde. 

a 

Cold  water. 

~h 



+ 

b 

Hot  water. 

ft 

y- 

-f- 

c 

AlcohoL 

s3^*&T 

~h 

-A 

d 

Ether. 

+- 

4- 

f_ 

Wrap  up  a  few  crystals  of  chloral  in  paper,  set  aside  and 
examine  them  after  a  few  days. 

In  what  ways  would  you  think  these  drugs  could  best  be 
prescribed  in  practice? 

II.  Put  a  few  crystals  of  chloral  in  a  test  tube  and  add 
a  small  amount  of  potassium  hydrate  solution  and  heat 
gently.  Note  the  odor  of  the  fumes.  What  is  formed  ? 


52  LABORATORY   GUIDE   IN 

CC13-CHO-H2O  +  KOH  =  H-COOK+  ? 
Chloral  hydrate.  Potassium  formate. 

III.  Select  four  frogs  of  about  equal  size  and  mark  each 
frog  in  some  way  so  that  it  can  be  recognized.  (This  is 
probably  done  most  easily  by  tying  a  piece  of  string  loosely 
around  one  of  the  legs  of  each  animal.)  Examine  them, 
making  a  record  of  the  heart  rate  and  respiration  in  each. 

Inject  into  the  anterior  lymph  sac  (page  15)  of  one  frog 
2.  mg.  of  chloral  dissolved  in  water ;  into  the  second,  inject  10 
mg.  chloral ;  into  the  third,  inject  20  mg. ;  and  into  the  fourth 
frog,  inject  50  mg.  These  amounts  may  be  obtained  most 
accurately  by  making  5  or  10  cc.  of  a  5%  solution  and  then 
diluting  small  quantities  ( i  cc . )  so  as  to  get  the  required 
doses. 

Make  a  record  of  the  time  when  each  injection  is  made. 
Replace  the  animals  under  the  bell  jar  and  observe.  Examine 
them  frequently  (especially  those  with  the  larger  doses)  as 
to  spontaneous  movements,  co-ordination,  reflexes,  respira- 
tion, heart  beat,  etc. 

Keep  a  careful  record  of  the  condition  of  each  frog  at  each 
examination. 

This  can  be   done  most  systematically,  not  only  in  this  experiment,  but  also 
in  those  to  follow,  by  keeping  a  record  of  the  time  when  each  examination  is 
made  and  the  conditions  found.     By  doing  this  it  also  gives  some  idea  of  the 
length  of  time  it  takes  the  drugs  to  act  when  given  in  different  sized  doses. 
Such  a  record  would  start  something  as  follows: 
Feb.    10,  '05 — Frog.      23   G.      String  on  right  front  leg. 
2:30 — Heart  56,  respiration  30  per  minute. 

2:32 — Injected  20  mg.  chloral  in  water  into  anterior  lymph  sac. 
2:35 — Frog  quiet,  apparently  normal;  reflexes,  normal;   heart,  54;  respir- 
ation, 28. 

2:40 — Frog  jumps  if  urged  strongly,  does  not  regain  sitting  position 
easily  when  it  alights;  has  difficulty  in  turning  over  when 
placed  on  back.  Reflexes  lessened;  heart,  40;  respiration,  15,  etc. 

Such  a  record  should  give  an  accurate  picture  of  the  con- 
dition of  the  animal  at  each  stage  of  the  drug's  action. 


EXPERIMENTAL  PHARMACOLOGY. 


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LABORATORY   GUIDE   IN 


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EXPERIMENTAL  PHARMACOLOGY.  55 

If  complete  paralysis  occurs  in  any  animal  determine 
whether  the  action  is  on  the  central  nervous  system  or  is 
peripheral. 

This  is  done  by  stimulating  over  the  lumbar  spine  with  the  electric  current 
and  if  the  muscles  of  the  leg  contract  the  paralysis  must  be  central;  if  they 
do  not  contract  see  whether  the  trouble  is  in  the  nerve  or  in  the  muscles  by 
stimulating  the  latter  directly  with  the  current. 

Also  in  an  animal  thus  paralyzed,  expose  the  heart 
(page  16)  and  examine  and  record  its  condition. 

With  the  knowledge  gained  of  the  physiology  of  the 
nervous  system  study  the  records  of  the  four  frogs  and  from 
the  symptoms  recorded  determine  the  points  of  action  of  the 
drug  in  the  varying  doses. 

IV.  Inject  into  the  anterior  lymph  sac  of  a  frog  I  cc.  of  a 
10%  solution  of  paraldehyde  in  water.  Compare  the  symp- 
toms shown  by  this  animal  with  those  manifested  by  the 
chloral  frogs  in  Exp.  III.  Pay  particular  attention  to  the 
condition  of  the  heart,  which  can  probably  be  sufficiently  well 
studied  (as  to  its  rate)  through  the  chest  wall  without  ex- 
posing it.  Keep  the  animal,  as  it  will  probably  recover. 

The  difference  in  action  of  the  two  drugs  upon  the  heart 
is  explained  by  the  difference  in  the  constitution  of  the  two 
molecules.  The  chloral  molecule  contains  chlorine  and  these 
chlorine  compounds  all  have  a  decided  action  on  the  heart 
muscle  which  is  absent  in  the  case  of  paraldehyde,  whose 
molecule,  C6H12O3,  contains  no  chlorine.  Compare  the 
action  of  chloroform  and  ether  on  the  heart  (page  132). 

Other  members  of  this  group  are,  Trional. 

Tetronal. 
Urethane. 
Chloretone. 
Chloralose,  etc. 


56  LABORATORY   GUIDE   IN 

Therapeutics.1 

The  chloral  group  of  drugs  depresses  the  central  nervous 
system.  For  this  purpose  it  would  be  used  in  cases  of  in- 
somnia and  of  nervous  excitement.  Chloral  and  its  allies  are 
very  valuable  also  in  delirium  and  in  various  forms  of  con- 
vulsions, as  for  instance,  those  due  to  poisoning  with  certain 
drugs  or  those  due  to  specific  diseases  (tetanus).  In  certain 
forms  of  insanity  (mania)  and  in  some  nervous  diseases 
with  increased  activity  of  the  brain  and  spinal  cord  (chorea 
or  St.  Vitus's  dance)  the  quieting  effects  of  these  drugs  are 
largely  made  use  of. 

On  account  of  its  action  on  the  heart  chloral  must  be  em- 
ployed with  care  in  persons  who  are  already  affected  with 
certain  forms  of  heart  disease  and  its  effect  on  this  organ 
must  also  be  watched  with  care  in  case  of  its  prolonged  use, 
or  better,  some  non-chlorine  containing  soporific  may  be 
used  in  its  stead. 


*At  the  conclusion  of  the  study  of  a  drug  group  a  short  discussion  of 
the  therapeutic  uses  of  the  drugs  contained  therein  will  be  given.  Such  dis- 
cussions are  not  intended  to  give  complete  resume  of  all  the  therapeutic  uses 
of  the  drugs  studied,  but  only  those  will  be  mentioned  as  would  be  indicated 
by  the  results  obtained  in  the  foregoing  experiments.  The  student  will  thus  be 
shown  how  the  knowledge  gained  by  the  science  of  Pharmacology  is  trans- 
ferred to  and  made  use  of  in  the  Therapeutic  art;  or,  in  other  words,  in 
rational  medicine. 


EXPERIMENTAL  PHARMACOLOGY.  57 


58  LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  59 


Percolation. 

The  U.  S.  Pharmacopoeia  gives  the  following  definition  of 
this  process:  "The  process  of  percolation  or  displacement 
consists  in  subjecting  a  substance  or  mixture  of  substances, 
in  powder,  contained  in  a  vessel  called  a  percolator,  to  the 
solvent  action  of  successive  portions  of  a  certain  menstruum 
in  such  a  manner  that  the  liquid,  as  it  traverses  the  powder 
in  its  descent  to  the  receiver,  shall  be  charged  with  -the  soluble 
portion  of  it,  and  pass  from  the  percolator  free  from  insol- 
uble matter." 

The  liquid  with  the  soluble  portions  of  the  drug  dissolved 
in  it  is  known  as  the  percolate. 

The  drug  to  be  percolated  is  usually  ground  to  a  fine 
powder  in  order  to  break  up  the  vegetable  cells,  etc.,  so  that 
the  menstruum  can  penetrate  it  easily,  and  the  menstruum  is 
allowed  to  pass  through  the  drug  only  very  slowly,  giving  it 
time  to  dissolve  the  soluble  constituents.  To  still  further  aid 
this  penetration,  a  common  plan  is  to  moisten  the  powder 
with  a  small  quantity  of  the  menstruum  and  to  allow  it  to  re- 
main tightly  covered  (macerate)  for  twenty- four  or  forty- 
eight  hours  before  beginning  the  percolation.  This  also 
gives  time  for  the  drug  to  swell,  as  is  likely  to  occur  in  some 
cases. 

Percolators1  are  usually  cylindrical,  or  conical,  and  vary 
in  size  with  the  amount  of  drug  to  be  exhausted.  The 
lower  orifice  is  to  be  closed  with  a  tightly  fitting  cork, 


JFor  all  purposes  in  this  course  the  cylindrical  percolator  of  the  following 
dimensions  is  large  enough:  Length,  28.2  cm.;  internal  diameter  at  top,  3.7 
cm. ;  capacity,  275  cc.  It  should  be  provided  with  an  ordinary  iron  stand  and 
ring,  the  latter  being  padded  with  cotton. 


60  LABORATORY   GUIDE  IN 

which  is  best  put  in  from  the  inside,  but  this  is  not  necessary 
with  the  small  percolators.  Through  this  cork  passes  a  small 
glass  tube  (5  mm.  in  diameter),  the  inner  end  of  the  tube 
being  flush  with  the  top  of  the  cork.  To  the  outer  end  of  the 
tube  is  fitted  a  piece  of  rubber  tubing.  One  method  of  regu- 
lating the  flow  of  the  percolate  is  to  have  this  tubing  of  suf- 
ficient length  to  reach  nearly  to  the  top  of  the  percolator  and 
to  have  the  free  end  fitted  with  another  glass  tube ;  the  latter 
can  be  raised  or  lowered  until  the  percolate  escapes  at  the 
desired  rate.  The  other  method,  which  probably  is  not  quite 
as  good,  is  to  have  the  rubber  tube  about  an  inch  long  and  to 
regulate  the  flow  by  means  of  a  small  screw  clamp. 

Having  fitted  the  cork  and  tubing  the  percolator  is  ready 
to  pack  if  the  drug  has  macerated  the  required  length  of 
time.  The  packing  is  done  by  means  of  a  plunger,  which  is 
made  by  taking  a  cork  with  a  diameter  about  half  as  great  as 
the  diameter  of  the  upper  opening  of  the  percolator  and 
sticking  into  it  a  glass  rod  which  is  a  little  longer  than  the 
depth  of  the  percolator. 

Place  in  the  bottom  of  the  percolator  a  small  piece  of  ab- 
sorbent cotton  and  press  it  loosely  in  place.  Only  a  small 
amount  of  cotton  should  be  employed,  as  its  only  use  is  to 
keep  the  powdered  drug  from  passing  down  and  clogging  up 
the  exit  tube.  Now  put  in  the  drug,  packing  it  uniformly  in 
place  with  the  plunger.  Do  not  use  much  force  or  the  drug 
will  be  so  tightly  packed  that  it  will  be  very  hard  for  the  men- 
struum to  pass  through.  Very  little  more  force  will  be  re- 
quired than  would  be  given  by  dropping  the  plunger  from  a 
height  of  a  few  inches.  Experience  together  with  a  knowl- 
edge of  the  physical  and  chemical  constitution  of  the  differ- 
ent drugs  can  alone  teach  how  much  force  to  employ.  When 
the  drug  is  in  place  cover  it  with  a  piece  of  filter  paper  cut 


EXPERIMENTAL  PHARMACOLOGY.  61 


62  LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  63 

so  as  to  roughly  fit  its  upper  surface  and  weight  it  down 
with  a  small  glass  stopper.  Pour  in  the  menstruum  to  the 
top  of  the  percolator  and  cover  the  latter  with  a  small  glass 
plate.  If  the  lower  orifice  is  open  the  menstruum  will  soon 
penetrate  through  the  drug  and  begin  to  drop  from  the  rub- 
ber tube.  In  cases  in  which  the  formula  prescribes  that  the 
drug  is  to  macerate  for  a  certain  length  of  time  in  the  perco- 
lator, the  clamp  is  closed,  or  if  a  long  tube  has  been  used  the 
end  is  raised  to  a  level  which  will  prevent  any  outflow.  At 
the  end  of  the  prescribed  time  allow  the  percolate  to  escape, 
receiving  it  in  a  bottle  or  flask.  Regulate  the  outflow  so  that 
it  shall  not  escape  faster  than  ten  or  fifteen  drops  in  a  minute. 
It  is  best  after  once  starting  percolating  to  keep  a 
layer  of  menstruum  above  the  surface  of  the  drug  so  as  to 
prevent  the  access  of  air  to  its  interstices.  If  the  latter  hap- 
pens the  drug  contracts  into  irregular  masses  leaving  fur- 
rows and  fissures,  and  when  more  menstruum  is  added  it 
will  naturally  flow  through  these  channels  and  will  not  pene- 
trate the  drug  evenly. 

The  U.  S.  P.  gives  general  directions  for  percolation  and 
also  in  the  cases  of  the  individual  drugs  specifies  the  size  of 
the  powder  and  the  number  of  grams  to  be  used  together 
with  the  amount  and  character  of  the  menstruum.  It  also 
gives  directions  as  to  the  length  of  time  the  drug  is  to  mac- 
erate before  percolation. 

Percolation  is  one  of  the  most  important  processes  em- 
ployed in  pharmacy,  as  by  it  are  prepared  extracts  (page 
179)  ;  fluid  extracts  (page  64)  ;  and  tinctures  (page  119). 
In  addition  it  is  one  of  the  steps  employed  in  the  isolation  of 
alkaloids. 

5-p 


64  LABORATORY  GUIDE  IN 


Nux  Vomica.  \ 

Nux  Vomica,  the  seeds  of  Strychnos  nux  vomica,  an  East 
Indian  tree,  contains  two  alkaloids  upon  which  the  activity  of 
the  drug  depends.  By  far  the  more  important  of  the  two  is 
Strychnine  which  is  present  in  from  .9  to  2.  per  cent;  the 
second  alkaloid,  Brucine,  is  present  in  slightly  smaller  quan- 
tities. In  action  the  two  alkaloids  resemble  each  other  very 
closely  excepting  that  strychnine  is  about  forty  times  as 
strong  as  brucine.  For  this  reason  the  activity  of  nux  vom- 
ica may  be  considered  as  dependent  upon  the  amount  of 
strychnine  present  and  in  the  following  experiments  it  alone 
will  be  considered. 

Examine  the  nux  vomica  bean  and  taste  a  little  of  the 
powdered  drug. 

I.  Place  a  small  amount  of  the  powdered  drug  in  a 
beaker  and  pour  upon  it  10  cc.  of  boiling  water  and  let  it 
stand  until  cold.     Set  this  watery  extract  (infusion)  aside 
and  use  it  in  the  experiments  given  below   (Exps.  IV,  V, 
VI).     Place  a  drop  of  the  infusion  on  the  tongue. 

II.  Prepare  the  fluid  extract  of  nux  vomica  in  the  follow- 
ing manner : 

Nux  vomica,  No.  60  powder — 50  grams. 

Alcohol  and  water  in  the  proportion  of  2 :  I. 

Moisten  the  powder  with  25  cc.  of  the  diluted  alcohol  and 
let  it  macerate  in  a  well  covered  vessel  in  a  warm  place  for 
48  hours.  Pack  it  in  a  percolator  (see  percolation,  page  59) 
and  gradually  pour  on  diluted  alcohol  until  the  drug  is  ex- 
hausted. Reserve  the  first  35  cc.  of  the  percolate  and  evap- 
orate the  remainder  over  a  water  bath  to  a  soft  extract.  Dis- 


EXPERIMENTAL  PHARMACOLOGY. 


66 


LABORATORY   GUIDU   IN 


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EXPERIMENTAL  PHARMACOLOGY.  67 

solve  this  extract  in  the  35  cc.  saved  and  make  the  total  bulk 
up  to  50  cc.  with  diluted  alcohol. 

Define  a  fluid  extract. 

III.  Reserve  5  cc.  of  the  fluid  extract  made  in  Exp.  II. 
and  from  the  remaining  45  cc.  isolate  the  strychnine  in  the 
following  manner : — 

Dilute  the  fluid  extract  with  about  its  own  volume  of  dis- 
tilled water  and  pour  it  into  a  separating  funnel  and  render 
it  alkaline  with  ammonia.  Add  about  25  cc.  of  a  mixture  of 
chloroform  and  ether  in  the  proportion  of  2:  I  and  shake 
gently  for  three  minutes  and  set  the  funnel  aside  to  allow  the 
chloroformic  solution  to  separate.  When  separation  has 
taken  place,  draw  the  chloroform  and  ether  off  into  a  beaker 
and  repeat  the  agitation  of  the  watery  extract,  using  a  fresh 
supply  of  chloroform  and  ether.  Draw  this  off  as  before 
into  the  beaker  with  the  first  solution.  The  watery  extract 
can  now  be  thrown  away.  Pour  the  chloroform  and  ether 
into  the  separating  funnel,  which  has  been  washed,  and  add 
about  25  cc.  of  water  and  enough  dilute  H.,SO4  to  make  the 
solution  distinctly  acid,  shake  gently  for  a  few  minutes  and 
then  separate  the  water  and  chloroform  as  before,  placing 
the  acidulated  water  in  a  flask.  Repeat  the  agitation  of  the 
chloroform  and  ether  with  fresh  acidulated  water,  adding 
the  latter  when  separation  has  taken  place,  to  the  first  lot  in 
the  flask.  (The  waste  chloroform  is  to  be  poured  into  a  large 
bottle  provided  for  it  and  later  redistilled.)  If  the  acidulated 
watery  solution  is  much  colored  it  should  be  shaken  in  the 
flask  with  charcoal  and  filtered.  Render  the  filtrate  dis- 
tinctly alkaline  with  ammonia  and  shake  'it  out  twice  as  be- 
fore in  the  separating  funnel,  with  small  quantities  of  the 
mixture  of  chloroform  and  ether.  When  this  has  been  done, 
pour  this  chloroformic  solution  into  a  glass  evaporating  dish, 


68  LABORATORY   GUIDE  IN 

add  to  it  2  or  3  drops  of  concentrated  hydrochloric  acid  ,and 
stir  it  well  with  a  glass  rod  or  put  the  mixture  in  a  small 
flask  and  shake  it  well.  The  solution  becomes  milky  from 
the  formation  of  strychnine  hydrochloride,  which  is  thrown 
out  of  solution  and  may  be  separated  from  the  chloroform 
by  allowing  the  latter  to  evaporate  spontaneously  or  by 
warming  it  gently  over  a  water  bath. 

CAUTION. — In  this  process  two  things  are  of  prime  im- 
portance in  determining  the  success  of  the  manipulations : — 
First,  to  agitate  sufficiently  the  chloroform  and  ammonia  or 
sulphuric  acid  solutions  to  allow  thorough  intermingling  of 
the  solutions.  Three  to  five  minutes  should  be  devoted  to 
each  shaking.  Second,  make  sure  that  the  correct  reaction 
is  present.  Test  with  litmus  paper  each  time  and  after 
shaking  for  a  minute  or  two,  test  again.  (See  Alkaloids, 
Exp.  No.  II.)  The  reaction  must  be  distinct  each  time. 

Explain  why  each  step  in  the  manipulation  was  carried 
out.  (Compare  Alkaloids,  Exp.  Nos.  I,  III,  VI.) 

The  crystallized  strychnine  hydrochloride  is  to  be  used  in 
tests  given  below.  (Exp.  Nos.  IX,  X,  XII.) 

IV.  To  a  few  cubic  centimeters  of  the  infusion  made  in 
Exp.  I  add  a  few  drops  of  ferric  chloride  solution.     The 
dark  color  of  the  mixture  is  due  to  the  tannic  acid  present 
in  the  nux  vomica  bean  uniting  with  the  iron  to  form  ferric 
tannate.      (To  a  dilute  solution  of  tannic  acid  add  ferric 
chloride.) 

V.  Boil  a  small  amount  of  Fehling  solution  and  add  3 
or  4  cc.  of  the  infusion  and  see  if  there  is  any  reduction. 

NOTE. — There  is  a  reducing  substance  found  in  the  nux  vomica  bean. 

VI.  Inject  I  cc.  of  the  infusion  into  the  lymph  sac  of  a 
frog,  and  replacing  the  animal  under  a  bell  jar,  observe  the 


EXPERIMENTAL  PHARMACOLOGY. 


69 


70 


LABORATORY   GUIDE   IN 

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EXPERIMENTAL  PHARMACOLOGY.  71 

effect.  Keep  full  notes.  If  any  convulsions  come  on, 
after  studying  them  carefully,  try  to  find  out  the  point  of 
action  of  the  drug  by  dividing  off  in  turn  the  cerebrum, 
optic  lobes,  and  finally  destroying  the  cord,  watching  the 
effect  of  each  operation  on  the  symptoms.  The  strychnine 
convulsion  is  "tonic"  as  distinguished  from  the  "clonic," 
which  is  caused  by  some  drugs  and  occurs  in  some  diseases. 
(Compare  Thujon,  page  79). 

VII.  Inject  into  the  lymph  sac  of  a  second  frog  o.i  cc.  of 
the  fluid  extract  (prepared  in  Exp.  II)  diluted  with  physio- 
logical salt  solution  to  make  a  convenient  amount.     In  this 
animal,  as  in  the  previous  experiment,  study  the  early  effects 
of  the  poison  before  convulsions  come  on.    Look  for  changes 
in  irritability,  in  reflexes,  etc.    If  convulsions  come  on,  place 
the  animal  in  a  beaker  which  has  been  partly  filled  with  water 
into  which  has  been  stirred  some  chloretone.     (The  latter 
drug  acts  in  this  case  as  a  local  anaesthetic.)     Does  it  alter 
the  character  of  the  convulsions  ? 

If  paralysis  should  occur  in  any  of  the  frogs  used  in  these 
experiments  see  if  it  is  due  to  central  or  peripheral  action,  as 
was  done  in  chloral,  Exp.  III. 

VIII.  Inject  into  a  small  frog  1/20  mg.  of  strychnine. 
When  convulsions  come  on  place  the  animal  under  a  bell  jar 
with  some  absorbent  cotton  upon  which  has  been  poured 
some  chloroform.     Observe  whether  any  change  takes  place 
in  the  convulsions.     If  they  become  lessened,  remove  the 
chloroform  from  the  bell  jar  and  allow  the  animal  to  recover 
from  its  effects.    Do  the  convulsions  return  ? 

Is  any  therapeutic  hint  contained  in  this  experiment?  In 
experiments  Nos.  VI,  VII  and  VIII,  it  was  noticed  that  the 
same  effects  were  induced  whether  the  poison  was  employed 
as  the  infusion,  as  the  fluid  extract,  or  as  the  pure  alkaloid. 


72  LABORATORY  GUIDE  IN 

IXa.  Put  one  or  two  crystals  of  the  strychnine  hydro- 
chloride,  prepared  in  Exp.  Ill,  on  a  porcelain  cover  and  add 
to  it  two  drops  of  concentrated  sulphuric  acid,  mixing  the 
two  with  a  clean  glass  stirring  rod.  Put  beside  the  strych- 
nine-sulphuric acid  mixture  on  the  cover  a  minute  crystal  of 
potassium  bichromate  and  with  the  glass  rod  (which  has 
been  washed  and  dried)  draw  the  bichromate  crystal  through 
the  strychnine  mixture.  Observe  the  changes  in  color.  This 
"oxidation-reaction"  or  "fading  purple  test"  for  strychnine 
is  most  delicate  and  characteristic.  It  is  said  to  be  capable 
of  detecting  i/2ooooth  of  a  grain  of  the  alkaloid.  The  order 
of  the  color  changes  is  very  important  in  making  the  test. 
The  purple,  which  should  first  appear,  changes  to  a  crimson, 
which  turns  to  a  cherry  red  that  is  fairly  persistent. 

IXb.  Carry  out  the  same  experiment  as  above,  using 
manganese  dioxide  instead  of  potassium  bichromate.  Com- 
pare the  results  with  Exp.  IXa. 

This  reaction  is  obtained  with  almost  any  oxidizing  agent, 
such  as  potassium  permanganate,  potassium  ferricyanide,  or 
lead  dioxide.  Manganese  dioxide  is  one  of  the  best  as  the 
changes  in  color  take  place  very  slowly,  in  some  cases  lasting 
some  minutes. 

If  much  brucine  is  present  in  the  preparation,  this  color 
test  is  much  interfered  with  and  it  may  be  impossible  to  get 
the  correct  changes. 

IXc.  After  testing  your  own  alkaloid  with  the  two  re- 
agents named,  make  the  same  tests  on  some  pure  strychnine 
which  will  be  furnished  you.  Compare  the  colors  with 
those  obtained  with  your  preparation. 

X.  Dissolve  the  rest  of  the  strychnine  hydrochloride 
(Exp.  Ill)  in  about  20  cc.  distilled  water  and  employ  it  in 
the  following  tests. 


EXPERIMENTAL   PHARMACOLOGY.  73 


74 


LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  75 

Inject  0.5  cc.  of  the  solution  into  a  frog.  Pith  (page  15) 
the  animal  and  hang  it  up  by  the  jaw  on  the  hook  for  re- 
flexes. When  the  strychnine  symptoms  appear  test  the 
animal's  reflexes  by  pinching  the  toe  and  by  dipping  the 
foot  in  acid.  How  do  the  reflexes  differ  from  those  in  a 
normal  animal? 

XI.1  The  object  of  the  experiment  is  to  apply  strychnine  to 
the  cervical  portion  of  the  cord  while  leaving  the  lumbar  cord 
unaffected.  Instead  of  pithing  a  frog  as  usual,  remove  the 
brain  entirely  by  cutting  off  the  head  with  a  pair  of  scissors. 
Put  the  lower  blade  in  the  mouth  and  the  upper  blade  over 
the  top  of  the  head,  incline  the  scissors  backward  slightly 
and  make  a  quick  cut.  Expose  the  heart  and  remove  it 
from  the  body  in  order  to  stop  the  circulation.  The  upper 
part  of  the  cord  will  be  exposed  by  the  cut  first  made  and 
a  drop  or  two  of  a  moderately  strong  strychnine  solution 
(0.1%)  is  to  be  applied  to  it  at  short  intervals.  Test  fre- 
quently the  reflexes  by  pinching  the  fore  and  hind  legs.  If 
the  experiment  is  successful  a  normal  reflex  should  be  ob- 
tained by  pinching  the  toes  of  the  hind  foot,  while  a  typical 
strychnine  convulsions  involving  the  entire  animal  should 
be  brought  on  by  irritation  of  the  fore  limbs. 

How  would  such  results  help  to  locate  the  point  of  strych- 
nine activity? 

(.See  Pharmacology  and  Therapeutics,  Cushny,  Edit.  Ill, 
page  200.) 

XII.  Put  the  remainder  of  the  solution  of  strychnine 
hydrochloride  into  5  test  tubes,  diluting  with  a  little  water 


1This  experiment  is  not  always  successful  when  carried  out  by  students  and 
may  be  omitted  if  thought  best.  However,  if  successful,  it  is  of  so  much  in- 
terest and  importance  in  locating  the  point  of  action  of  the  drug  as  to  merit 
a  place  in  these  notes. 


76 


LABORATORY   GUIDE  IN 


if  necessary  and  test  the  solutions  with  the  various  alkaloidal 
reagents. 


Reagent 

Precipitate? 

Limit  of  delicacy  of 
reagent 

a 

Tannic  acid. 

>+    sy^r&^f- 

b 
c 

Picric  acid.                             ' 
Iodine  in  potassium  iodide. 

^tx£^-W~ 

1-20,000 
1-100,000 

d 

Mercury-potassium  iodide. 

/-  ^1/2^6; 

1-150,000 

e 

Phosphotungstic  acid. 

+    */^£ 

1-200,000 

XIII.  With  a  hypodermic  syringe  inject  subcutaneously 
into  a  rabbit  I  mg.  of  strychnine  sulphate.  Watch  the  effect 
on  reflexes,  etc.  If  convulsions  come  on  control  them  with 
a  few  drops  of  chloroform  on  cotton  held  close  to  the  ani- 
mal's nose. 

Compare  the  convulsions  in  this  animal  with  those  seen  in 
the  frog.  Note  the  efficiency  of  chloroform  in  controlling 
the  convulsions  of  strychnine. 


Therapeutic 

On  account  of  their  intensely  bitter  taste,  preparations  of 
nux  vomica  are  employed  in  cases  of  loss  of  appetite  and 
malnutrition.  By  their  action  on  the  taste  organs  they 
probably  increase  reflexly  the  secretion  of  the  gastric  juice 
and  thus  aid  digestion. 

Strychnine  is  largely  used  as  a  stimulant  to  the  central 
nervous  system,  being  useful  in  various  conditions  in  which 
depression  of  the  brain  or  cord  is  found.  It  is  employed  in 
shock  or  collapse,  and  in  failure  of  the  respiratory  or  of 
the  vaso-motor  center.  By  stimulating  the  activity  of  the 
spinal  cord  to  a  slight  extent  it  increases  the  "tone"  of  the 
muscles  and  of  the  blood  vessels  whose  muscular  coats  are 


EXPERIMENTAL  PHARMACOLOGY.  77 


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EXPERIMENTAL   PHARMACOLOGY.  79 

controlled  by  the  spinal  centers.     Perhaps  more  than  any 
other  drug  it  deserves  the  name  "tonic." 

Thujon. 

Thujon  is  the  poisonous  principle  derived  from  the  volatile 

oils  of  sage,  tansy  and  absinthe.    It  is  an  isomere  of  camphor 

(C10H16O)  and  may  be  taken  as  an  example  of  that  series, 

as  it  acts  in  the  body  in  a  manner  closely  resembling  that 

tdrug. 

I.  Place  a  cat  in  a  cat-box  (page  12)  and  pass  a  tube 
through  the  mouth  gag  into  the  stomach  and  inject  by  means 
of  a  pipette  about  i  cc.  of  thujon.  The  end  of  the  pipette 
containing  the  thujon  is  inserted  into  the  opening  of  the 
stomach  tube  and  the  drug  blown  in  and  afterward  washed 
down  with  some  water.  The  stomach  tube  and  gag  are  now 
removed  and  the  animal  is  freed  and  placed  in  a  cage. 
Compare  the  symptoms  with  those  caused  by  strychnine  in 
the  rabbit.  It  will  be  seen  that  the  convulsions  caused  by 
thujon  are  of  a  distinctly  different  type  from  those  following 
a  poisonous  dose  of  strychnine.  Those  caused  by  the  latter 
drug  are  tonic  in  type  and  are  due  to  an  action  on  the 
spinal  cord.  On  the  other  hand,  the  thujon  (and  camphor) 
convulsions  are  "clonic"  and  are  caused  by  stimulation  of 
some  unknown  part  of  the  cerebrum  or  lower  part  of  the 
brain. 

Therapeutic  uses** 

Thujon  is  not  employed  in  therapeutics.    Its  pharmacolog- 
ical action  would  indicate  that  it  would  be  of  value  as  a 
stimulant  to  the  central  nervous  system  and  especially  to 
6-p 


80  LABORATORY   GUIDE  IN 

the  respiratory  center.     As  such  it  would  be  of  value  in 
shock  and  collapse  and  perhaps  in  narcotic  poisoning. 
Camphor  has  long  been  used  in  shock  and  collapse. 


Opium. 

Opium  is  the  inspissated  juice  obtained  from  the  fruit- 
capsules  of  the  white  poppy  (Papaver  somniferum).  The 
capsules  are  incised  while  yet  unripe  and  the  milky  juice 
allowed  to  exude  and  dry  by  spontaneous  evaporation  until 
it  becomes  a  brownish  gummy  mass. 

The  poppy  is  grown  in  Turkey,  Asia  Minor,  Persia,  India 
and  China,  but  that  obtained  from  Smyrna  and  Turkey  is  by 
far  the  most  valuable.  Persian  and  Indian  opium  is  im- 
ported mainly  as  sources  of  the  opium  alkaloids  and  the 
Chinese  opium  is  consumed  entirely  in  that  country. 

Opium  contains  about  twenty  alkaloids  of  which  Morphine 
and  Codeine  are  the  most  important,  and,  indeed,  it  may  be 
said  that  the  value  of  opium  depends  upon  the  per  cent  of 
morphine  which  it  contains.  The  U.  S.  Pharmacopoeia 
specifies  that  the  crude  opium  when  moist  shall  contain  not 
less  than  9  per  cent  of  crystallizable  morphine,  while  the 
dried  product  must  assay  from  13  to  15  per  cent.  Opium 
from  the  different  localities  varies  exceedingly  in  its  mor- 
phine content. 

Morphine  was  discovered  in  1817  by  Seitiirner,  of  Eim- 
beck,  Germany,  and  was  named  by  him  morphium.  It  exists 
in  the  opium  principally  in  combination  with  meconic  acid, 
although  some  may  be  present  as  a  lactate  or  a  sulphate,  both 
of  these  acids  being  found  in  the  crude  drug.  Meconic 
acid  is  of  most  importance  as  it  is  strictly  characteristic  of 


EXPERIMENTAL   PHARMACOLOGY.  81 


82  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  83 

opium  and  its  preparations,  and  its  positive  detection  is  proof 
of  the  presence  of  some  opium  preparation. 

In  their  solubilities  morphine  and  codeine  differ  widely 
from  the  other  alkaloids,  for  morphine  is  almost  insoluble 
in  chloroform  and  only  very  slightly  soluble  in  ether 
(1-6100);  codeine,  on  the  other  hand,  is  more  soluble  in 
water  than  any  other  alkaloid. 

Morphine  Reactions. 

I.  Put  a  few  crystals  of  morphine  on  a  porcelain  cover 
and  add  a  drop  or  two  of  pure  concentrated  sulphuric  acid 
and  mix  the  two  with  a  clean  glass  rod.     No  color  should 
be  formed  if  the  morphine  is  absolutely  pure,  but  occasionally 
a  faint  pink  tint  is  seen.    Heat  the  mixture  very  gently.    At 
about  150°  a  dirty  green  or  rose-red  color  is  developed,  and 
on  still  further  heating,  the  solution  becomes  almost  black. 
Allow  it  to  cool  and  add  some  water,  when  a  greenish  blue 
color  appears   which   changes  to  green   when   ammonia  is 
added  in  excess.  / 

II.  Put  a  few  crystals  of  morphine  on  a  cover  and  mix 
them  with  a  drop  or  two  of  concentrated  sulphuric  acid  as  in 
Exp.  I.     To  this  add  a  drop  of  distilled  water  which  will 
heat  the  mixture.     Now  treat  with  a  drop  of  concentrated 
nitric  acid,  which  should  give  a  rose-red  color,  changing  to 
brown. 

(This  reaction  for  morphine  is  very  delicate,  being  well 
shown  by  o.oi  milligram  of  the  alkaloid.) 

III.  To  a  drop  of  concentrated  nitric  acid  on  a  porcelain 
cover  add  a  minute  amount  of  morphine.     An  orange-red 
color  is  obtained,  which  changes  to  yellow  on  heating. 

IV.  (a)     Mix  a  small  amount  of  morphine  with  about 
twice  its  weight  of  cane  sugar.    By  means  of  a  glass  rod  add 


/^ 


84  LABORATORY  GUIDE  IN 

a  drop  of  concentrated  sulphuric  acid.  This  should  produce 
a  purple  color,  changing  gradually  to  blood-red  and  brown- 
ish-red and  becoming  an  olive-brown  on  dilution  with  water. 
(b)  The  above  test  can  be  modified  as  follows:  —  To  a 
solution  of  morphine  add  cane  sugar  until  the  solution  is 
saturated.  Now  pour  this  solution  carefully  down  the  side 
of  a  second  test  tube  in  the  bottom  of  which  is  concentrated 
sulphuric  acid.  By  inclining  the  second  tube  the  solution 
can  be  poured  in  so  that  no  mixture  of  the  two  fluids  takes 
place  and  a  purple  or  rose-red  ring  will  form  at  their  junc- 
tion. This  ring  becomes  more  intense  on  standing. 

V.  Dissolve  some  morphine  sulphate  in  a  little  water  and 
add  it  to  a  solution  of  iodic  acid.    What  color  does  it  turn  ? 
Add  to  the  resulting  mixture  some  thin  starch  paste.   What 
was  the  first  color  due  to? 

NOTE.  —  In  order  tp  color  the  starch  paste  blue  morphine  must  be  present  in 
the  strength  of  i  to  1000.  If  no  blue  is  obtained  mix  the  solution  well  and 
pour  down  the  side  of  the  test  tube  some  very  dilute  ammonia.  A  double  ring 
will  be  seea  at  the  junction  of  the  two  fluids;  it  will  be  blue  below  and  brown 
above. 

VI.  To  a  few  cubic  centimeters  of  water  add  some  mor- 
phine sulphate  and  to  this  solution  add  a  few  drops  of  ferric 
chloride  solution.     A  deep  blue  color  is  obtained. 

I.     Meconic  acid  reaction.     To  a  cubic  centimeter  of 
tmcture  of  opium  diluted  with  a  little  water  add  two  drops 
—  of  ferric  chloride  solution.     What  color  does  the  solution 
become?     If  the  color  is  obscured  by  a  precipitate,  filter 
the  solution. 

VIII.  Inject  into  the  anterior  lymph  sac  of  a  frog  25  mg. 
of  morphine  sulphate.  Replace  the  animal  under  the  bell 
jar  and  observe.  Examine  it  from  time  to  time,  comparing 
the  effects  with  those  caused  by  chloral  or  strychnine. 


EXPERIMENTAL   PHARMACOLOGY.  8S 


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86  LABORATORY   GUIDE  IN 


EXPERIMENTAL  PHARMACOWGY.  87 

NOTE.— This  experiment  (VIII)  should  be  begun  early  in  the  laboratory  ses- 
sion as  it  sometimes  takes  two  or  three  hours  for  the  full  effects  to  appear. 

Among  the  alkaloids  found  in  opium  which  are  of  less 
importance  than  those  studied  is  thebaine,  which  resembles 
strychnine  very  closely  in  its  pharmacological  action.  In 
fact,  many  of  the  alkaloids  of  opium  may  be  arranged  in  a 
series  with  the  depressant,  morphine,  at  one  end  of  the  series 
and  thebaine  at  the  opposite  end. 

IX.  Inject  subcutaneously  into  a  dog  50  mg.  of  morphine 
sulphate  and  observe  the  effects. 

X.  Inject  30  to  40  mg.  of  morphine  sulphate  subcutane- 
ously into  a  cat.    Put  the  animal  in  a  wire  cage  and  compare 
the  effects  with  those  seen  in  the  dog. 

The  peculiar  action  of  morphine  on  the  cat  is  not  confined 
to  that  animal,  but  is  found  in  all  the  members  of  the  cat 
tribe,  and  also  in  the  horse  and  ass. 

Examine  a.  crude  opium, 

b.  powdered  opium, 

c.  extract  of  opium, 

d.,  tincture  of  opium    (Laudanum), 

e.  camphorated  tincture  of  opium   (Paregoric), 

f.  powdered  opium  and  ipecac  (Dover's  powder) . 

g.  and  codeine. 

Note  the  odor  of  all  the  opium  preparations  and  taste  d. 
e,  and  f. 

Therapeutics. 

The  symptoms  induced  by  opium  in  man  resemble  more 
closely  those  in  the  dog  than  those  in  the  other  animals.  It 
depresses  the  brain,  and  for  this  reason  is  used  in  cases  of 
sleeplessness  when  this  condition  results  from  pain,  because 
in  depressing  the  cerebrum  it  acts  on  the  pain  centers,  for 


88  LABORATORY  GUIDE  IN 

which  it  seems  to  have  a  special  affinity.  In  the  dog  the 
respiration  was  slowed  in  the  later  stages  of  the  action  from 
depression  of  the  respiratory  center,  and  opium  prepara- 
tions are  often  employed  in  cough  mixtures  to  allay  exces- 
sive irritability  of  this  part  of  the  medulla.  The -numerous 
other  purposes  for  which  opium  and  morphine  are  used 
cannot  be  illustrated  experimentally. 

Curara. 

Curara,  the  South  American  arrow  poison,  varies  in  com- 
position with  the  different  localities  from  which  it  is  ob- 
tained. The  best  known  product  comes  from  Guiana  and  is 
prepared  from  the  bark  of  Strychnos  toxifera,  a  tree  native 
to  that  country.  Curara  as  obtained  in  commerce  consists  of 
dark  brown,  shining,  brittle  masses  which  contain  vegetable 
extracts  in  addition  to  two  alkaloids,  tubocurarine  and 
curine.  The  characteristic  action  of  the  drug  is  dependent 
principally  upon  the  first  alkaloid  mentioned,  the  latter  hav- 
ing quite  a  different  action  and  being  much  weaker. 

Not  only  do  solutions  of  curara1  deteriorate  on  keeping, 
but,  what  is  of  greater  importance,  the  crude  drug,  as  ob- 
tained in  the  market,  varies  greatly  in  strength,  many  of  the 
preparations  being  entirely  inert.  There  is  no  chemical 
method  by  which  the  activity  of  curara  can  be  estimated; 
the  only  test  is  pharmacological.  Each  solution  to  be  used 
has  to  be  "standardized"  by  tests  on  animals  and  the  dose 
necessary  to  produce  characteristic  effects  ascertained. 

I.  Inject  into  a  frog  a  poisonous  dose  of  a  standardized 
solution  of  curara.  Study  the  effect  on  spontaneous  move- 


1The   curara    solution    should    be    made    up    in    physiological    salt   solution    to 
,'hich   some  thymol   has  been   added. 


EXPERIMENTAL  PHARMACOLOGY.  89 


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EXPERIMENTAL   PHARMACOLOGY.  91 

ments,  reflexes,  ability  to  turn  over  when  placed  on  its  back, 
and  length  of  time  leg  muscles  will  remain  contracted  when 
stimulated  by  applying  the  tetanizing  current  to  the  lumbar 
spine.  If  paralysis  should  occur  see  if  it  is  due  to  central  or 
peripheral  action  (see  Soporifics,  Exp.  Ill,  page  55).  This 
animal  is  to  be  replaced  under  the  bell  jar  and  kept  moist,  as 
it  will  probably  recover  in  a  few  days. 

II.  Pith  a  frog,  tie  it  on  the  frog  board  and  expose  the 
sciatic  nerve  and  artery  along  the  back  of  the  thigh  by  a 
short  longitudinal  incision  through  the  skin,  and  careful 
separation  of  the  muscles  with  the  dissecting  needle.  Sepa- 
rate the  sciatic  nerve  from  the  surrounding  tissues  along  a 
short  distance  of  its  course  and  then  pass  a  thread  under 
the  nerve  and  tie  it  tightly  around  the  rest  of  the  thigh  as 
high  up  as  possible. 

Free  the  animal,  inject  curara  as  in  I,  and  from  time  to 
time  compare  the  reflexes  in  the  hind  legs.  Stimulate  the 
lumbar  cord  with  the  electric  current  and  note  any  differences 
between  the  two  sides.  See  whether  the  reflexes  remain 
equal  in  the  two  legs  and  whether  cross  reflexes  can  be 
obtained. 

If  the  experiment  is  successful  the  operated  leg  should  not 
be  affected  by  the  poison  and  should  respond  to  electrical 
stimulation.  In  such  a  case  cut  off  the  leg  just  above  the 
knee,  leaving  attached  to  it  the  sciatic  nerve  which  has  been 
carefully  separated  from  the  thigh  muscles  and  cut  toward 
the  upper  part  of  its  course.  Holding  the  leg  preparation 
by  the  severed  end  of  the  femur  lay  the  hanging  sciatic 
nerve  for  a  few  moments  in  the  curara  solution.  Remove  it 
and  stimulate  it  with  the  electric  current.  Does  curara  affect 
the  nerve  trunk?  Dip  the  muscle  in  the  solution  and  after 
a  few  moments  stimulate  the  sciatic  nerve  again.  Is  there 


9:  LABORATORY   GUIDE  IN 

any  change?    If  so,  test  the  activity  of  the  muscles  by  direct 
stimulation.    Where  does  curara  act  ? 

Examine  the  heart  of  the  animal  in  Exp.  II. 

Examine  Curara. 

Therapeutics. 

The  main  importance  of  curara  is  in  physiological  experi- 
ments. It  has  been  suggested,  and  in  some  few  cases  used, 
to  allay  the  convulsions  due  to  tetanus  or  to  strychnine 
poisoning.  It  is  not  recommended  for  such  conditions  as 
its  action  is  too  uncertain  and  too  dangerous. 

Nicotine. 

The  alkaloid,  nicotine,  is  obtained  from  the  dried  leaf  of 
the  tobacco  plant  (Nicotianum  Tabacum)  in  which  it  is 
found  to  the  extent  of  from  2  to  8  per  cent.  The  pure  alka- 
loid is  a  colorless,  oily  fluid,  which,  on  exposure  to  air,  be- 
comes yellowish  or  brown.  It  has  a  strong,  unpleasant  odor 
resembling  somewhat  that  of  tobacco.  When  tobacco  is 
smoked,  only  a  small  amount  of  the  alkaloid  appears  in  the 
smoke,  the  larger  part  is  decomposed  in  the  process  and  it 
may  be  that  some  of  the  effects  of  smoking  are  due  to  the 
absorption  of  these  decomposition  products  rather  than  to 
the  nicotine  itself. 

In  an  aqueous  solution  the  alkaloid  gives  a  strong  alkaline 
reaction  and  it  unites  with  various  acids  to  form  salts. 

I.  Inject  i  cc.  of  nicotine  chloride  solution1  into  the 
anterior  lymph  sac  of  a  frog.  Observe  the  effects,  noting 
especially  the  posture  of  the  animal  in  the  early  stages  of 


1The  nicotine  chloride  solution  is  formed  by  neutralizing  a  small  quantity  ot 
nicotine  with  1-20  normal  hydrochloric  acid.  Each  cubic  centimeter  will  then 
contain  about  4  mgs.  of  nicotine. 


EXPERIMENTAL   PHARMACOLOGY.  93 


94 


LABORATORY   GUIDE   IN 


,  v^        _ 


HXPHRIMHNTAL  PHARMACOLOGY.  95 

poisoning  as  this  is  very  characteristic  of  nicotine  action. 
In  addition,  watch  for  any  twitching  of  the  muscles,  and  if 
paralysis  should  come  on  see  whether  it  is  due  to  central  or 
peripheral  action. 

The  action  of  nicotine  upon  the  sympathetic  nervous  sys- 
tem is  studied  later  (pages  187,  195). 

Therapeutics. 
Nicotine  is  not  employed  in  therapeutics. 

Veratrine. 

Veratrine  is  an  alkaloid  prepared  from  the  seeds  of  Asa- 
groea  officinalis.  It  is  a  grayish  powder  which  causes  very 
marked  irritation  when  even  a  minute  quantity  is  inhaled. 

The  alkaloid  is  not  of  great  medical  importance  but 
is  of  pharmacological  interest,  owing  to  a  peculiar  action  it 
possesses  which  can  be  best  studied  in  cold-blood  ani- 
mals, although  the  same  effect  may  be  elicited  in  mammals. 

I.  Inject  i  mg.  of  veratrine  sulphate  (i  cc.  of  o.i  %  sol.) 
into  a  frog.  Watch  and  describe  the  effects.  Observe  the 
awkwardness  of  its  movements  which  appears  in  a  few  min- 
utes. When  the  clumsiness  is  well  developed  locate  the 
point  at  which  the  drug  acts  by  removing  the  cerebrum, 
optic  lobes,  and  finally  destroying  the  spinal  cord  if  the 
symptoms  do  not  disappear  earlier.  Stimulate  the  sciatic 
plexus  with  single  shocks  from  the  induction  coil,  comparing 
the  results  with  those  seen  in  a  normal  frog.  Finally,  stimu- 
late the  muscle  directly,  using  single  shocks.  At  what  point 
does  the  drug  act? 
7-p 


96 


LABORATORY  GUIDE  IN 


Therapeutics. 

The  phenomena  seen  in  this  experiment  give  no  hint  as  to 
the  use  of  the  drug  in  therapeutics;  it  is  employed  for  ex- 
ternal application,  and  internally  to  a  limited  extent  for  its 
effects  on  the  circulation. 


Caffeine. 

Caffeine  differs  from  the  other  alkaloids  in  the  fact  that  it 
is  contained  in  a  number  of  plants  belonging  to  different 
species.  It  is  obtained  from  the  berry  of  the  coffee  (Coffea 
Arabica)  ;  from  the  leaves  of  tea  (Thea  Chinensis),  besides 
being  present  in  the  kola  nut  of  Central  Africa;  in  the 
Paraguay  tea  and  Guarana  paste  of  South  America  and  in 
the  Apalache  tea  of  Virginia  and  Carolina. 

In  some  of  these  plants  it  is  associated  with  two  closely 
related  alkaloids,  Theobromine  and  Theophyllin,  which  re- 
semble caffeine  very  much  in  some  of  their  actions.  All 
three  alkaloids  are  derivatives  of  xanthine,  which  in  turn 
is  closely  related  to  uric  acid. 

Caffeine  is  a  very  feeble  base  and  forms  salts  only  with 
difficulty  and  these  salts,  as  a  rule,  are  very  unstable,  being 
easily  decomposed,  even  by  water. 

I.    Test  the  solubility  of  caffeine  in 


Reagent. 

Solubility. 

/ 
J^c-tx^ 

a 

Water,  cold. 

+                     rf    A        A 
/  tQ  V 
/_  // 

b 
c 
d 

hot. 
Alcohol. 
Chloroform. 

+  If' 

^        (ah 

e^u 

^^ 

EXPERIMENTAL  PHARMACOLOGY 


<0~ 


/? 


98 


LABORATORY   GUIDti   IN 


EXPERIMENTAL  PHARMACOLOGY. 


99 


II.    Apply  the  various  alkaloidal  reagents  to  a  i%  solu- 
of  caffeine. 


Reagent. 

Precipitation? 

a 

Tannic  acid. 

^s5n!^ 

b 

Picric  acid. 

—  • 

c 

Iodine  in  potassium  iodide. 

-  — 

d 

Mercury-potassium  iodide. 

— 

e 

Phosphotungstic  acid. 

+  <»L^ 

NOTE. — In  its  reaction   to  c  and   d  caffeine   differs    from  all   other  alkaloids 
except  theobromine  and  colchicine.  \| 

III.  Murexom  test.   To  a  few  crystals  of  caffeine  on  the 
porcelain  cover  add  a  drop  of  concentrated  nitric  acid.  Hold 
the  cover  with  a  pair  of  forceps  some  distance  above  the 
flame  of  a  Bunsen  burner  and  thus  gently  evaporate  the 
mixture  to  dryness.    Allow  the  residue  to  come  in  contact 
with  strong  ammonia  fumes  and  note  the  color  obtained. 

This  reaction  is  given  by  the  members  of  the  xanthine 
series  and  is  practically  identical  with  that  given  by  uric 
acid. 

IV.  Inject  5  mg.  of  caffeine  (i  cc.  0.5  %  sol.)  into  a 
frog.     Study  and  describe  the  symptoms  comparing  them 
with  those  induced  by  strychnine  and  veratrine.    See  whether 
the  peculiar  change  in  the  muscles  is  due  to  an  action  on 
the    nervous    system    by    destroying    its    various    parts    as 
was   done   with   veratrine.     Excise   a   small   piece   of  the 
affected  muscle  and  tease  it  in  salt  solution  on  a  microscopic 
slide.    Examine  it  under  the  microscope,  using  the  low  power 
lens,  and  compare  its  appearance  with  normal  muscle  pre- 
pared in  the  same  way.     (For  an  explanation  of  the  changes 
consult  Pharmacology  and  Therapeutics,  Ed.  Ill,  page  248.) 


100  LABORATORY   GUIDE  IN 

The  action  of  caffeine  on  the  heart  and  on  the  kidney  will 
be  studied  later  (pages  139,  159,  172). 

Examine  the  Coffee  bean  (Coffea  Arabica)  ; 

Tea  leaves  (Thea  Chinensis)  ; 

Caffeine  citrate; 

Theobromine. 

Therapeutic  Uses. 

Caffeine  is  largely  employed  as  a  central  nervous  stimu- 
lant. The  stimulant  action  on  the  highest  parts  of  the  brain 
is  more  noticeable  in  man  than  in  the  frog  as  the  cerebrum 
in  the  latter  is  not  as  highly  differentiated  and  the  principal 
symptoms  are  due  to  cord  action.  In  man,  however,  caffeine 
(usually  taken  in  the  form  of  coffee  or  tea)  stimulates  the 
cerebrum  and  the  medulla  and,  to  a  less  extent,  the  spinal 
cord.  It  is  of  great  use  in  cerebral  depression,  such  as  is 
found  in  shock  and  collapse.  In  narcotic  poisoning,  also,  its 
action  on  the  medullary  centers  is  especially  valuable.  Both 
caffeine  and  theobromine  are  largely  used  for  their  stimu- 
lant action  on  the  kidneys  by  which  they  increase  the  urine. 
They  are,  therefore,  of  service  in  removing  abnormal  col- 
lections of  fluid  from  the  body,  such  as  oedema  or  ascites, 
whether  of  cardiac  or  renal  origin. 

Cocaine. 

Cocaine  is  the  principal  alkaloid  derived  from  the  leaves 
of  Erythroxylon  coca,  a  shrub  which  is  a  native  of  Peru  and 
Bolivia  and  of  other  parts  of  South  America.  It  is  very 
unstable,  being  quite  prone  to  undergo  chemical  change,  so 
that  preparations  of  the  drug  frequently  contain  the  products 
of  its  decomposition.  For  this  reason  there  are  no  very 


EXPERIMENTAL   PHARMACOLOGY.  101 

^U^ZL. 


102 


LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  103 

delicate  or  distinctive  chemical  tests  by  which  the  drug  may 
be  identified.  Being  an  alkaloid,  it  is  thrown  out  of  solution 
by  the  various  precipitates  of  the  group,  and  in  its  behavior 
to  solvents  it  resembles  in  general  the  other  alkaloids. 

Cocaine  has  an  important  local  action  upon  the  termina- 
tions of  sensory  nerves,  which  makes  it  of  very  great  use  in 
medicine. 

I.  Take  a  piece  of  filter  paper  2  or  3  cm.  square  and 
moisten  it  with  a  few  drops  of  a  4%  cocaine  hydrochloride 
solution  and  place  it  upon  the  tip  of  the  tongue.  Retain  it 
in  this  position  for  two  or  three  minutes  and  then  examine 
the  condition  of  that  part  of  the  tongue  exposed  to  the  drug, 
testing  it  especially  as  to  the  sense  of  touch,  temperature,  and 
taste.  Contrast  its  effect  on  the  taste  of  sugar  and  of  nux 
vomica. 

Therapeutic  Uses. 

Cocaine  is  most  largely  employed  in  medicine  for  its  action 
in  paralyzing  the  sensory  nerve  endings  causing  a  local 
anaesthesia,  permitting  of  operations  being  performed  with- 
out pain. 

Cinchona. 

In  the  plants  belonging  to  the  cinchona  family  are  a  num- 
ber of  species  the  bark  of  which  yields  some  twenty-five  alka- 
loids. Quinine  is  by  far  the  most  important  of  these.  The 
Cinchonacae  are  natives  of  South  America  and,  as  far  as 
is  known,  were  introduced  into  medicine  about  1630. 

Quinine  may  be  regarded  as  a  typical  alkaloid  and  its  be- 
havior towards  various  solvents  and  precipitants  has  already 
been  examined  (page  24). 


10*  LABORATORY   GUIDE   IN 

In  addition  to  those  properties,  which  it  shares  in  common 
with  the  other  alkaloids,  it  shows  a  few  reactions  of  its  own 
which  are  more  or  less  characteristic. 

Taste  quinine  and  quinine  hydrochloride. 

I.  Weigh  out  o.i  G.  quinine,  the  same  amount  of  quinine 
sulphate,  and  of  quinine  hydrochloride  and  put  them  sepa- 
rately in  three  test  tubes.    Add  10  cc.  of  water  to  each  test 
tube  and  shake.      Which  is  the  most  soluble?      If  either 
preparation  does  not  dissolve  in  this  amount  of  water,  add 
more  water,  a  definite  amount  each  time  (25  cc.)  and  see  if 
you  can  ascertain  approximately  the  solubility  of  the  differ- 
ent preparations. 

II.  To  o.i  G.  quinine  sulphate  in  a  test  tube,  add  about 
10  cc.  distilled  water  and  about  I  cc.  of  dilute  sulphuric  acid. 
Note  the  change  in  solubility  compared  with  I  and  describe 
the  solution  formed.     Solutions  showing  this  peculiar  fluor- 
escence are  also   formed  by  a  few  of  the  other  cinchona 
alkaloids. 

III.  Make  a  solution  of  quinine  sulphate  in  a  test  tube, 
adding  a  drop  or  two  of  sulphuric  acid  if  necessary.    Add 
very  carefully  potassium  hydroxide  to  make  the  solution  as 
nearly  neutral  as  possible  (not  alkaline!).    Add  a  few  drops 
of  chlorine  water  and  after  shaking  add  ammonia  to  excess. 
Observe  the  greenish  coloration  of  the  solution  or  the  green 
precipitate  which  will  be  formed  if  enough  quinine  is  present. 

This  reaction  is  of  importance,  as  it  can  be  employed  in 
testing  for  quinine  in  the  urine  in  cases  in  which  it  is  im- 
portant to  know  that  the  drug  is  being  absorbed. 

IV.  Take  0.3  to  0.5  G.  quinine  sulphate  and  test  a  sample 
of  the  urine  passed  within  the  next  twelve  hours  for  the 
presence  of  quinine  according  to  the  above  method.     Note 


EXPERIMENTAL   PHARMACOLOGY.  105 

^^^-^-t-^^tS^o^^^-^'^^57 


106  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  107 

whether  any  symptoms  on  part  of  the  eyes  or  ears  are  ob- 
served. 

Examine  Cinchona  bark. 

Tincture  of  cinchona. 

Compound  tincture  of  cinchona. 

Therapeutic  Uses. 

The  tinctures  of  cinchona  are  largely  employed  in  medi- 
cine on  account  of  their  bitter  taste;  they  are  given  espe- 
cially in  cases  of  lack  of  appetite.  The  other  therapeutic 
uses  of  cinchona  and  its  alkaloids  cannot  be  demonstrated 
experimentally  in  the  laboratory. 


108  LABORATORY   GUIDE   IN 


Emulsions. 

Emulsions  are  aqueous  preparations  in  which  oils,  resins 
or  insoluble  powders  are  suspended  by  means  of  gummy 
substances  (emulsifiers). 

The  object  of  emulsification  is  to  render  substances  in- 
soluble in  water  freely  miscible  in  that  fluid,  so  that  any  dis- 
agreeable taste  may  be  more  easily  disguised  and  the  ab- 
sorption of  the  oil  rendered  more  easy.  In  nature,  some 
emulsions  exist  already  formed,  as  for  example,  milk,  the 
juice  of  plants,  the  yolk  of  eggs,  etc. 

There  are  a  large  number  of  substances  which  can  be  used 
as  emulsifiers,  the  most  important  being  acacia.  Condensed 
milk  and  yolk  of  egg  are  superior  to  acacia  for  this  purpose, 
but  possess  the  disadvantage  of  becoming  stale. 

In  some  substances,  as  for  instance  in  gum  resins,  the  gum 
needed  to  form  an  emulsion  is  found  present  with  the  resin. 
In  the  seeds  of  some  plants  albuminous  substances  are  found 
and  these  act  as  emulsifiers  of  the  oils  present  in  the  seed, 
rendering  the  addition  of  gums  unnecessary. 

The  various  types  of  emulsions  are  prepared  according  to 
the  following  directions : 

I.     Seed  emulsion  prepared  from  bitter  almonds. 

Take  four  or  five  bitter  almonds  and  remove  their  skins. 
This  may  be  done  in  two  ways.  They  may  be  dropped  for 
a  few  moments  in  very  hot  water  and  then  in  cold,  when  the 
skins  can  be  slipped  off.  The  danger  with  this  method  is 
that  the  boiling  water  may  destroy  the  ferment,  emulsin, 
which  aids  in  the  emulsification.  To  avoid  this,  the  seeds 


EXPERIMENTAL   PHARMACOLOGY.  109 


LABORATORY   GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  Ill 

may  be  soaked  in  lukewarm  water  for  half  an  hour,  or  until 
their  skins  are  loosened. 

Break  up  the  seeds  and  put  them  in  a  clean  mortar  and 
add  a  small  quantity  of  water  (one  part  to  ten  of  seeds) 
and  with  the  pestle  rub  the  seeds  up  into  a  thick,  creamy 
paste  which  shall  be  free  from  lumps.  More  water  can  be 
added  now,  gradually,  until  a  milky  solution  is  formed, 
which  may  be  strained  if  necessary. 

II.  Emulsion  of  a  gum  resin. 

Prepare  100  cc.  of  the  emulsion  of  asafcetida  according  to 
the  U.  S.  P. 

III.  Emulsion  of  a  fixed  oil. 

Among  the  various  methods  employed  in  making  emul- 
sions, that  known  as  the  Continental  is  probably  the  most 
satisfactory,  and  is  to  be  used  in  the  'following  experiment. 

Prepare  60  cc.  of  a  50%  emulsion  of  castor  oil. 

In  this  method  a  primary  emulsion  or  nucleus  is  first 
formed  with  certain  proportions  of  the  ingredients  and  this 
nucleus  can  then  be  diluted  with  water  without  the  fear  that 
separation  will  occur. 

The  proportion  by  weight  is  as  follows : 

Oil : 4 

Water   2 

Gum  (powdered  acacia) I 

Using  this  formula,  calculate  the  amount  of  each  ingredi- 
ent needed  and  weigh  them  out  carefully  in  clean  vessels. 
Pour  the  oil  into  a  clean,  dry  mortar  and  add  the  acacia,  and 
with  the  pestle  stir  the  two  together  rapidly  until  a  uniform 
mixture  is  obtained.  (This  must  be  done  quickly,  as  other- 
wise the  oil  and  acacia  remain  in  contact  too  long  and  the 
latter  tends  to  become  hard  and  insoluble.  The  object  is 
merely  to  get  each  particle  of  gum  incased  in  oil.)  To  the 
8-p 


112  LABORATORY   GUIDE   IN 

oil-acacia  mixture  add  all  the  water  (as  determined  by  the 
above  formula)  and  triturate  the  mixture  rapidly  until  it 
becomes  a  thick,  white,  creamy  emulsion.  In  the  trituration, 
which  should  take  from  3  to  5  minutes,  no  pressure  is  needed, 
but  a  rapid  motion  is  very  important.  The  pestle  should  be 
held  loosely  between  the  thumb  and  first  two  or  three  fingers 
and  the  motion  given  not  only  from  the  shoulder  and  elbow, 
but  also  from  the  wrist  and  fingers.  It  is  best  to  move  the 
pestle  always  in  one  direction,  but  it  is  not  essential  to  do  so. 

When  the  nucleus  is  complete  it  can  be  poured  into  a 
bottle  and  water  added  to  make  up  the  required  bulk.  It 
should  then  be  well  shaken.  . 

Such  an  emulsion  may  separate  after  some  hours  into  two 
layers  without  showing  any  separated  oil.  Shaking  will 
restore  the  uniform  appearance.  Such  separation  is  en- 
tirely analogous  to  that  which  occurs  in  milk  when  the 
cream  rises  to  the  surface. 

Emulsions  can  be  flavored  by  various  agencies,  such  as 
the  volatile  oils,  but,  as  a  general  rule,  some  of  the  less 
powerful  flavors  will  disguise  the  taste  of  the  oil  better,  as 
for  instance,  liquorice  extract,  coffee,  vanilla,  or  chocolate. 
They  may  -be  sweetened  by  small  quantities  of  sugar  or 
saccharin. 

Acids,  alcohol,  or  glycerin,  if  added  in  any  except  the 
smallest  quantities,  cause  the  emulsion  to  separate  (break). 

Pills. 

Pills  are  spherical  or  ovoid  masses  of  medicinal  substances 
of  a  size  convenient  for  swallowing.  In  weight  they  should 
not  exceed  0.3  G.,  the  average  being  about  0.2  G. 

This  method  of  administering  solid  medicinal  substances 


EXPERIMENTAL  PHARMACOLOGY.  113 


114  LABORATORY   GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  115 

is  perhaps  more  common  than  any  other,  partly  on  account 
of  convenience  and  also  because  substances  which  have  a 
disagreeable  taste  may  be  given  most  agreeably  in  this  way. 
Drugs  which  are  corrosive  or  are  very  deliquescent  or  which 
have  to  be  given  in  very  large  doses  should  not  be  given  in 
pills.  Those  which  are  best  suited  for  administration  in  this 
form  are  powders  and  extracts. 

The  first  step  in  pill  making  is  to  form  the  pill  mass,  which 
should  be  of  doughy  consistency  but  not  sticky.  The  mass 
consists  of  two  parts;  the  active  ingredients  which  are  to 
be  incorporated  in  the  pill  and  some  substance  (the  ex- 
cipient)  which  will  give  the  proper  degree  of  consistency 
and  adhesiveness.  The  excipient  will  vary  with  the  char- 
acter of  the  active  ingredients.  Some  of  the  powdered  ex- 
tracts only  require  water  to  give  them  the  proper  consis- 
tency, while  dilute  alcohol  may  be  needed  in  other  extracts. 
Some  fluid  preparations  may  be  made  into  pills  by  using  an 
inert  powder  and  on  the  other  hand  a  non-adhesive  powder 
may  be  made  into  a  pill  mass  by  the  use  of  soft  extracts 
or  by  the  use  of  such  an  excipient  as  a  combination 
of  glycerin  and  tragacanth  (glycerite  of  tragacanth).  The 
choice  of  an  excipient  is  very  important,  as  it  largely  de- 
termines the  usefulness  of  pills  if  they  are  kept  for  some 
time.  For  example,  a  pill  made  with  acacia  as  excipient 
may  become  so  hard  that  it  will  pass  through  the  body  un- 
changed, while  with  other  excipients  the  pill  may  dry  out 
and  crumble  to  pieces  after  they  have  been  made  for  a  short 
time.  The  formation  of  a  pill  mass  may  be  learned  best  by 
filling  the  following  prescription,  which  calls  for  ten  pills 
to  be  made  from  a  gram  of  extract  of  gentian. 

IJ.     Extract!  Gentianae,  i. 
Divide  in  pilulas,  No.  X. 


116  LABORATORY  GUIDE  IN 

Counterpoise  two  slips  of  paper  on  the  scale  pans  and 
weigh  out  the  correct  amount  of  extract,  which  is  then  trans- 
ferred to  a  mortar.  (If  some  of  the  extract  sticks  to  the 
piece  of  paper  moisten  the  opposite  side  of  the  paper  with 
water  and  wait  a  few  moments,  after  which  the  remains  of 
the  extract  may  be  easily  scraped  off.) 

Make  the  extract  into  a  pill  mass  of  the  character  described 
above.  If  it  is  too  hard  add  a  little  water  or  dilute  alcohol ; 
if  too  soft  add  a  small  quantity  of  a  powder,  such  as  starch  or 
the  powdered  extract  of  liquorice.  Triturate  the  mass  thor- 
oughly so  as  to  get  the  two  substances  uniformly  distributed 
through  each  other.  When  the  mass  is  completed,  work  it  up 
with  the  hands  into  a  short  cylinder  and  transfer  it  to  the 
pill  slab,  over  which  has  been  scattered  a  small  quantity  of 
a  dusting  powder,  such  as  lycopodium.  Roll  out  the  mass 
with  the  wooden  roller  into  an  even  cylinder  of  the  right 
length  to  correspond  with  the  scale  on  the  slab  (in  this  case 
roll  out  to  10),  taking  care  that  the  ends  of  the  cylindrical 
mass  are  "squared  up."  With  the  cylinder  over  the  scale, 
cut  it  with  the  spatula  into  the  required  number  of  pieces, 
taking  care  that  the  spatula  is  always  held  at  right  angles  to 
the  mass.  The  little  sections  of  the  pill  mass  are  now  to  be 
moulded  with  the  thumb  and  fingers  into  spherical  shape 
and  then  placed  in  the  pill  box  with  a  small  amount  of  lyco- 
podium and  shaken  about  so  that  they  may  be  completely 
covered  by  powder  which  will  prevent  them  sticking  to- 
gether. Discard  the  excess  of  powder. 

To  mask  the:  taste  of  the  pills  they  are  usually  coated  with 
some  substance  such  as  gelatin,  sugar,  chocolate,  etc. 

Examine  the  pills  after  a  day  or  two  and  see  if  they  have 
retained  their  shape  or  if  they  have  flattened  out.  Weigh 
five  pills  and  see  how  nearly  they  correspond  in  weight. 


EXPERIMENTAL   PHARMACOLOGY.  117 


118 


LABORATORY  GUIDE  IN 


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EXPERIMENTAL   PHARMACOLOGY.  119 


The  Digitalis  Series. 

This  series  embraces  a  large  number  of  drugs  which  re- 
semble each  other  in  their  pharmacological  action.  While 
they  act  on  various  organs  in  the  body,  the  cardiac  action  is 
characteristic  of  the  group  and  distinguishes  it  from  all 
others.  The  members  of  the  series  are  derived  from  plants 
widely  distributed  in  nature  and  as  widely  separated  in  their 
botanical  relationships. 

The  most  important  is  digitalis,  which  is  obtained  from 
Digitalis  purpurea  (purple  fox  glove)  ;  strophanthus  from 
Strophanthus  hispidus;  and  squills  from  Scilla  maritima. 

I.  Prepare  the  tincture  of  digitalis  according  to  the  U.  S. 
P.,  using  50  G.  of  the  powdered  drug  and  the  proportionate 
amount  of  dilute  alcohol. 

NOTE. — For  the  strength  of  "dilute  alcohol"  consult  the  U.  S.  P.  under 
"Alcohol  Dilutum." 

Define  a  tincture. 

This  tincture  is  to  be  used  in  the  following  experiments: 

II.  To  5  cc.  of  the  tincture  add  15  cc.  of  distilled  water. 
Is  a  precipitate  formed?     Cork  and  set  aside  the  mixture 
and  examine  it  after  a  few  days. 

III.  Examine   the  tincture   for   the   presence   of  tannic 
acid  by  adding  ferric  chloride. 

IV.  To  5  cc.  of  the  tincture  add  3  cc.  of  tannic  acid  solu- 
tion and  let  it  stand  for  a  time.    Is  a  precipitate  formed  ? 

X  x,    '  * 

(Digitalis  contains  glucosides  some  of  which  are  precipi- 
tated by  tannic  acid.) 

V.  To  5  cc:  of  the  tincture  add  a  few  drops  of  mercury- 


120  LABORATORY   GUIDE  IN 

potassium  iodide.    Is  a  precipitate  formed?    What  does,  the 

^ 

result  indicate? 

VI.  Evaporate  over  a  water  bath  10  cc.  of  the  tincture  to 
about  one-half  its  volume  and  then  make  up  the  concentrated 
tincture  to  the  original  bulk  by  adding  physiological  salt 
solution. 

Now,  pith  a  frog  and  tie  it  on  a  board.  Expose  the  heart 
and  inject  into  a  lymph  sac  I  cc.  of  the  modified  tincture. 
Record  all  changes  in  the  heart. 

Describe  any  change  in  the  rate  of  the  organ  and  also  any 
differences  in  the  size  of  the  various  chambers  or  in  their 
manner  of  contracting  or  dilating. 

VII.  Pharmacological  assay  of  tincture1  of  digitalis. 
Digitalis  preparations  vary  considerably  in  their  strength 

and  it  is  impossible  to  standardize  them  by  any  chemical 
means  such  as  can  be  employed  in  the  cases  of  those  drugs 
whose  activity  depends  upon  alkaloids  which  may  be  isolated 
in  pure  form  and  weighed. 

It  is  very  hard  to  isolate  the  glucosides  in  pure  form  as 
they  are  likely  to  break  up,  making  the  results  obtained  very 
unreliable.  For  this  reason  a  pharmacological  estimation  of 
the  strength  of  a  preparation  is  employed  as  follows : — 

Evaporate  10  cc.  of  the  tincture  over  a  water  bath  to  about 
half  volume  and  then  make  it  up  to  the  original  volume  by 
the  addition  of  physiological  salt  solution.  Select  3  frogs  of 
about  the  same  size,  weigh  them  carefully  and  mark  them 
with  string  so  as  to  be  able  to  identify  them.  Into  the  an- 
terior lymph  sac  of  one  of  the  frogs  inject  by  means  of  a 
glass  pipette  0.3  cc.  of  the  modified  tincture  diluted  with 


xThe  assay  of  the  tincture  may  be  performed  by  one-half  the  class  while  the 
infusion  of  digitalis   (page  124)    is  assayed  by  the  remainder. 


EXPERIMENTAL   PHARMACOLOGY. 


121 


122  LABORATORY   GUIDE  IN 


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EXPERIMENTAL   PHARMACOLOGY.  123 

salt  solution;  into  frog  No.  2,  inject  0.2  cc. ;  and  into  the 
third  frog,  o.i  cc.  (These  small  quantities  may  be  most  ac- 
curately measured  by  diluting  the  modified  tincture  still  more 
with  salt  solution  and  then  injecting  the  proportionately 
larger  dose.) 

Keep  a  record  of  the  time  when  each  injection  is  made. 
Replace  the  frogs  under  the  bell  jar  and  at  the  end  of  one 
hour  from  the  time  of  each  injection  examine  the  condition 
of  each  frog's  heart.  Pith  the  animal  if  necessary,  tie  it  on 
the  frog  board  and  expose  the  heart  according  to  the  usual 
method.  To  have  a  correct  "end  reaction"  the  heart  should 
have  just  ceased  beating  with  the  ventricle  in  systole  and  the 
two  auricles  markedly  distended  with  blood.  (Compare 
Exp.  VI.)  If  all  three  hearts  are  still  beating  when  they  are 
examined  the  doses  were  too  small  and  must  be  increased  in 
fresh  frogs  of  corresponding  weight.  If,  on  the  other  hand, 
the  frogs  have  been  dead  for  some  time,  the  doses  were  too 
large  and  must  be  made  smaller  in  other  frogs. 

For  moderate  sized  frogs  (15-20  G.)  the  toxic  dose  will 
probably  be  between  the  limits  given,  viz.:  0.3  cc.  to  o.i  cc. 
By  varying  the  quantity  given  the  standard  for  a  frog  of  a 
certain  weight  may  be  ascertained. 

Calculate  the  dose  of  your  tincture  necessary  to  kill  a 
standard  frog  of  20  G.  weight  in  an  hour.  Why  did  you 
evaporate  the  tincture  to  one-half  its  volume? 

The  method  of  assay  described  is  of  considerable  im- 
portance as  it  is  employed  by  pharmaceutical  firms  for  the 
standardization  of  many  cardiac  remedies. 

The  remainder  of  the  tincture  is  to  be  saved  to  be  used  in 
subsequent  experiments  on  blood  pressure,  etc. 

The  effects  of  digitalis  on  the  mammalian  heart  and  on  the 
arterial  blood  pressure  are  studied  later.  (Exps.,  pages  159 


124  LABORATORY   GUIDE   IN 

and  151).  Its  effect  on  the  excretion  of  urine  (diuretic 
action)  may  be  studied  in  the  rabbit  (note,  page  175),  and 
the  vascular  changes  in  the  intestinal  and  renal  vessels  ac- 
cording to  directions  given  on  pages  227  and  232). 

Infusion  of  Digitalis. 

Another  preparation  of  digitalis  which  is  largely  used  in 
some  parts  of  the  world  in  place  of  the  tincture  and  which 
it  resembles  in  most  of  its  actions  is  the  Infusion. 

I.  Prepare  the  infusion  of  digitalis  according  to  the  U. 
S.  P. 

Define  an  infusion. 

II.  Test  the  infusion  for  glucosides  with  tannic  acid. 

III.  Fill  a  test  tube  about  half  full  with  the  infusion, 
place  the  thumb  over  the  mouth  of  the  tube  and  shake  it  up 
thoroughly.    Note  the  soap-like  character  of  the  foam.    The 
infusion  contains  a  glucoside,  digitonin,  which  possesses  the 
property  of  forming  a  frothy  solution  and  of  holding  in- 
soluble bodies  in  suspension. 

IV.  Pith  a  frog,  tie  it  on  the  board  and  expose  the  heart 
as  usual.    Inject  into  a  lymph  sac  I  cc.  of  the  infusion.  Com- 
pare the  effects  with  those  noticed  in  the  experiments  in 
which  the  tincture  was  employed. 

V.  See  foot  note,  page  120. 

Assay  the  infusion  of  digitalis  by  the  method  described 
under  ''Tincture"  (Exp.  VII,  page  120),  employing,  instead 
of  0.3  cc.,  0.2  cc.  and  o.i  cc.  of  the  tincture,  2  cc.,  1.5  cc. 
and  i  cc.  of  the  infusion.  Also,  in  making  the  assay,  it  will 
not  be  necessary  to  evaporate  the  preparation  as  was  done 
with  the  tincture.  Why? 

VI.  Save  about  100  cc.  of  the  infusion  and  examine  it 


EXPERIMENTAL   PHARMACOLOGY.  125 


126 


LABORATORY   GUIDE   IN 


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EXPERIMENTAL   PHARMACOLOGY.  127 

from  time  to  time  and  note  any  changes  taking  place  in  it. 
Examine  digitalis  leaves  (Digitalis  purpurse)  ; 

strophanthus  seeds  ( Strophanthus  hispidus)  ; 

and  squills,  the  sliced  bulb  (Scilla  maritima). 
Also  examine  the  following  preparations  : — 

Tincture  of  strophanthus ; 

Tincture  of  squills ; 

Syrup  of  squills. 

Therapeutic  Uses. 

Digitalis  and  its  allies  are  employed  principally  in  certain 
affections  of  the  heart.  When  a  valve  of  the  heart  is  diseased 
so  that  it  does  not  perform  its  functions  properly,  the  heart 
has  an  increased  amount  of  work  to  do.  In  most  cases  its 
walls  increase  in  thidmess  (hypertrophy)  to  allow  of  the 
extra  work  being  done,  in  other  words  the  heart  becomes 
'''compensated."  BuH£  an  unusual  demand  is  made  on  the 
heart  before  it  has  time  to  hypertrophy,  the  walls  stretch  on 
account  of  their  weakness,  and  the  heart  is  said  to  become 
"dilated."  When,  fhis^eccurs  the  contractions  are  weak  and 
imperfect  and  the  result  is  that  the  circulation  becomes 
slowed  and  the  blood  tends  to  cotK^ct  on  the  venous  side  of 
the  vascular  system,  resulting  in  a  passive  congestion  of  the 
organs,  and  in  marked  cases  a  generalized  oedema  of  the 
body  may  follow. 

In  such  conditions  digitalis  is  of  the  greatest  value*  As 
was  seen  in  its  effect  on  the  dog's  heart,  it  increases  the 
strength  of  the  contractions  and  in  some  cases  lessens  the 
extent  of  dilatation.  The  heart  begins  to  pump  out  more 
blood  into  the  arteries  raising  the  arterial  pressure  and  thus 
supplying  more  nutrition  to  the  body  cells.  One  of  the  first 
organs  to  be  benefited  is  the  heart  itself  through  its  coronary 
9-p 


123  LABORATORY  GUIDE  IN 

arteries.  From  its  improved  blood  supply  it  becomes  stronger 
and  may  be  able  to  hypertrophy  and  thus  carry  on  its  in- 
creased work  without  the  further  use  of  digitalis.  The  im- 
proved circulation  results  in  a  disappearance  of  the  conges- 
tion of  the  organs  and  of  the  oedema. 

The  removal  of  the  oedema  is  also  aided  by  the  diuretic 
action  of  the  drugs  belonging  to  this  series ;  this  appears  to 
be  due  mainly  to  the  cardiac  action  (cardiac  diuretic). 

Aconite. 

Several  alkaloids  are  derived  from  plants  belonging  to1  the 
Aconitum  genus,  and  among  them  Aconitine  is  the  most  im- 
portant. It  is  obtained  from  the  root  of  the  monk's  hood 
(Aconitum  Napellus). 

The  most  important  preparation  containing  aconitine  is  the 
tincture  of  aconite,  the  alkaloids  being  very  rarely  used  as 
different  preparations  vary  considerably  in  activity,  owing  to 
the  fact  that  aconitine  decomposes  with  great  readiness. 

I.  To  2  cc.  of  the  tincture  of  aconite  add  an  equal  amount 
of  water.    Try  to  dissolve  the  precipitate  formed  by  adding 
some  95%  alcohol.     The  precipitate  in  this  case  consists  of 
resinous  bodies  (extractives)  which  are  thrown  out  of  solu- 
tion by  the  water  but  which  are  redissolved  by  the  alcohol. 

II.  Add  6  drops  of  tincture  of  aconite  to  about  I  cc.  of 
distilled  water  in  a  clean  beaker.     Take  the  solution  in  the 
mouth  and  retain  it  there  without  swallowing  for  about  a 
minute,  after  which  it  may  be  ejected  and  the  mouth  rinsed 
with  water.     Observe  whether  any  unusual  sensation  comes 
on  in  from  five  to  ten  minutes  in  that  part  of  the  mouth  with 
which  the  aconite  solution  has  come  in  contact. 

These  symptoms,  which  are  due  to  stimulation  of  the  sen- 


EXPERIMENTAL  PHARMACOLOGY.  129 


130  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  131 

sory  nerve  endings,  are  characteristic  of  two  drugs,  aconite 
and  veratrine. 

III.  Pith  a  frog,  tie  it  on  the  board  and  expose  the  heart 
by  the  usual  method  (page  16).  Inject  into  a  lymph  sac  3 
drops  of  tincture  of  aconite  and  keep  a  record  of  all  changes 
taking  place  in  the  organ,  noting  not  alone  changes  in  rate, 
but  also  in  the  character  of  the  contractions. 

Examine  the  root  of  Aconitum  Napellus. 

The  effect  of  aconite  on  the  mammalian  heart  is  studied 
later.  (Page  139). 


132  LABORATORY   GUIDE   IN 


The  Effect  of  Drugs  on  the  Frog's  Heart. 

Pith  a  frog,  tie  it  on  a  board  and  expose  the  heart  as 
usual.  Dissect  out  the  vagus  nerve  and  stimulate  it  with  a 
weak  tetanizing  current  from  the  induction  coil.  To  find  the 
vagus  nerve  in  the  frog :  expose  the  heart  as  usual,  split  the 
pectoral  girdle  entirely  and  draw  the  anterior  limbs  well 
apart.  Carefully  remove  the  skin  and  connective  tissue  in 
the  neck  and  two  large  nerves  (9"  and  12")  will  be  seen  on 
either  side  passing  forward.  Trace  these  backward  and  they 
will  be  found  to  take  a  turn  outward  to  enter  the  base  of  the 
skull.  The  vagus  in  this  position  lies  between  them,  running 
parallel  to  them  in  this  part  of  their  course. 

Describe  the  changes  in  the  heart  caused  by  vagus  stimu- 
lation. 

NOTE. — In  a  certain  number  of  cases  the  vagus  in  the  frog  is  not  active  and 
it  will  therefore  be  impossible  to  affect  the  heart  by  its  electrical  stimulation. 

In  this  frog  or  in  a  second  one  if  necessary,  examine  the 
action  of  chloroform  by  putting  the  animal,  with  the  heart 
exposed,  under  the  bell  jar  in  which  there  is  a  piece  of  ab- 
sorbent cotton  soaked  in  chloroform. 

Record  all  changes  seen  in  the  heart,  whether  changes  in 
rate  or  in  the  condition  of  the  various  chambers. 

If  the  effects  become  very  marked,  remove  the  animal  from 
the  bell  jar  and  allow  the  heart  to  recover.  Keep  it  moist 
by  dropping  on  it  a  small  amount  of  physiological  salt  solu- 
tion. 

When  the  organ  has  recovered  repeat  the  experiment,  us- 
i'ng  ether  instead  of  chloroform. 

Compare  the  actions  of  the  two  drugs.     (See  Angesthet- 


EXPERIMENTAL  PHARMACOLOGY.  133 


134  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  135 

ics,  page  204).  Also  compare  the  action  of  chloral  on  the 
heart.  (Page  55). 

After  the  heart  has  returned  once  more  to  normal,  put  a 
drop  of  a  pilocarpine  (or  muscarine)  solution  on  it  and  note 
any  change  in  rate.  After  a  few  minutes  add  a  few  more 
drops  of  the  same  solution  and  note  any  further  change. 
Compare  the  appearance  of  the  heart  with  its  condition  under 
vagus  stimulation.  If  the  heart  should  stop  contracting,  put 
two  drops  of  an  atropine  solution  on  the  organ  and  if  neces- 
sary, after  a  few  minutes  make  a  second  or  third  application 
of  atropine. 

If  the  heart  begins  to  beat  once  more  apply  pilocarpine 
solution  again  and  see  if  it  will  stop  it.  Also  try  stimulation 
of  the  vagus  nerve.  If  pilocarpine  will  not  stop  the  heart, 
try  chloroform  by  exposing  the  organ  to  the  fumes  as  before. 

From  your  knowledge  of  the  anatomy  and  physiology  of 
the  vagus  nerve  try  to  account  for  the  effects  induced  by  the 
drugs  as  seen  above,  considering  them  in  the  following 
order,  pilocarpine,  atropine,  pilocarpine  and  chloroform. 

The  effects  of  digitalis,  aconite,  chloral  and  £araldehyde 
on  the  he'art  are  studied  elsewhere.  (Pages  120,  131  and  55). 

The  Effect  of  Drugs  on  the  Rabbit's  Heart. 

Anaesthetize  a  small  rabbit  or  cat  as  described  elsewhere, 
(page  n).  When  anaesthesia  is  complete  tie  the  animal  on 
the  operating  board  and  insert  a  tracheal  tube  (page  23), 
and  a  venous  cannula  in  the  jugular  vein  (page  19).  Find 
one  of  the  vagus  nerves,  separate  it  gently  from  the  other 
structures  in  the  carotid  sheath  and  pass  a' thread  under  it. 
Tie  the  thread  and  cut  the  nerve  above  the  ligature  so  as  to 
allow  of  stimulation  of  the  peripheral  end. 


136 


LABORATORY  GUIDE  IN 


Make  an  incision  down  the  thorax  in  the  median  line  as 
far  as  the  end  of  the  sternum  and  deepen  the  cut  to  the  bone, 
taking  care  to  keep  exactly  in  the  middle  line.  Start  arti- 
ficial respiration  with  the  bellows,  after  which  split  the  ster- 
num longitudinally  by  strong  scissors  and  draw  its  two 
halves  apart  by  fish  hooks,  securing  them  to  the  sides  of  the 
board.  The  heart  is  then  seen  within  the  pericardium,  which 
is  opened  with  scissors. 

Examine  the  contractions  of  the  normal  heart  and  then 
stimulate  the  vagus  with  both  weak  and  strong  currents,  ob- 
serving the  changes  in  the  beat. 

In  a  small  evaporating  dish  place  5  mg.  of  pilocarpine  ( I 
cc.  of  0.5  %  solution)  diluted  with  salt  solution  to  a  con- 
venient bulk  (about  5  cc.).  In  a  second  dish  place  I  mg.  of 
atropine  sulphate  (i  cc.  of  o.i  %  solution)  diluted  with  salt 
solution. 

Inject  the  pilocarpine  solution  into  the  jugular  vein  (see 
precautions  page  20)  by  means  of  a  hard  rubber  syringe, 
noting  the  effect  on  the  heart.  If  the  latter  is  very  much 


FIG.   No.    5. 


EXPERIMENTAL  PHARMACOLOGY. 


(S.(S 


138 


LABORATORY   GUIDU   IN 


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EXPERIMENTAL  PHARMACOLOGY.  139 

affected  inject  the  atropine  solution  at  once,  noting  also  its 
effect. 

After  the  atropine  injection  stimulate  the  vagus  as  before, 
noting  any  difference  in  its  effect  on  the  heart. 

Insert  in  the  air  tube  a  bottle  (Fig.  5)  containing  chloro- 
form, using  great  care  not  to  give  too  much.  Describe  the 
changes  in  the  heart.  Remove  the  chloroform  and  allow  the 
heart  to  recover  and  repeat  the  experiment,  using  ether  in- 
stead of  chloroform.  Which  is  the  more  powerful? 

After  the  heart  has  recovered  from  the  ether  inject  into  the 
jugular  vein,  by  means  of  a  syringe,  the  following  drugs 
in  the  order  named.  Describe  fully  the  changes  in  the  heart 
induced  by  the  various  drugs  in  these  doses : 

a.  Caffeine,  25  mg.  dissolved  in  2  cc.  of  hot  water. 

b.  Extract  of  the  suprarenal  gland.1 

c.  Calcium  chloride,  20  mg.  dissolved  in  2  cc.  of  water. 

d.  Potassium  chloride,  20  mg.  dissolved  in  2  cc.  of  water. 

e.  Aconite,  3  drops  of  the  tincture. 

NOTE. — Do  not  inject  one  drug  until  the  action  of  the  previous  drug  has 
passed  off. 

The  Effect  of  Drugs  on  the  Turtle's  Heart. 

Destroy  the  brain  of  a  turtle  by  a  blow  with  a  hammer,  and 
turning  the  animal  on  its  back,  tie  it  on  an  operating  board. 
Draw  the  head  out  so  as  to  put  the  neck  on  the  stretch  and 
fasten  it  in  this  position  by  means  of  a  nail  driven  through 
it.  Remove  the  lower  shell  (plastron),  cutting  the  lateral 
attachments  with  bone  forceps  or  a  saw ;  after  cutting  the 
hard  parts  the  shell  can  be  raised  slightly  and  the  underlying 
soft  parts  severed  with  a  scalpel,  taking  care  to  cut  toward  the 
shell.  Carefully  cut  away  the  skin  and  fascia  near  the  base 


1Inject    s    cc.    of    a    solution    made    by    adding    i    cc.    of    adrenalin    chloride 
(i-iooo)   to  50  cc.  physiological  salt  solution.      (See  page  213). 


140 


LABORATORY  GUIDH  IN 


of  the  neck,  and  if  the  coracoids  (G)  and  clavicles  (F)  are 
in  the  way  they  may  be  cut  back  with  bone  forceps  and  a 


FIG.  No.    6.   Turtle  with  plastron  removed  and  vagi  dissected  out.     A.  Coraco-hyoid- 

eus  muscle,  B.  Retrahens  capitis  collique  muscle  (right  muscle  is  cut  across), 

C.   Sympathetic  nerve,  D.  Vagus,  E.  Carotid  artery,  F.  Clavicle, 

cut  end,  G.  Coracoid,  cut  end,  H.  Heart. 

scalpel.    Large  white  muscles  (B,  B,),  the  long  retractors  of 
the  head  (Retrahens  capitis  collique)  will  now  be  exposed, 


EXPERIMENTAL   PHARMACOLOGY.  141 


</.     6/ 


142  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  143 

running  back  from  the  head  and  placed  deeply  in  the  neck  on 
either  side  of  the  median  line.  Two  other  large  white 
muscles  (coraco-hyoideus,  A)  will  also  be  seen  passing  on 
either  side  from  the  front  legs  forward  toward  the  median 
line  of  the  neck.  Two  nerves  (C,  D)  may  be  found  emerg- 
ing from  under  the  long  retractors  just  posterior  to  the  point 
where  the  coraco-hyoidei  unite  in  the  median  line.  In  the 
upper  part  of  their  course  they  lie  internal  to  the  long  re- 
tractors, then  wind  around  the  muscle  to  reach  its  upper  sur- 
face (in  this  position  of  the  animal)  and  pass  down  the  neck, 
lying  on  the  muscle.  One  of  these  nerves  is  the  vagus  (D), 
which  must  be  identified  by  gently  separating  it  from  the 
surrounding  structures  and  stimulating  it  with  the  electric 
current,  watching  the  effect  on  the  heart.  When  it  is  found, 
place  a  ligature  around  it. 

Expose  the  heart  by  cutting  away  the  pericardium. 
Thread  a  small  curved  needle  with  about  six  or  eight  inches 
of  medium  sized  thread  and  take  a  stitch  in  either  side  of  the 
ventricle,  trying  to  include  between  them  the  area  of  greatest 
contraction.  Tie  the  thread  in  the  heart  with  a  loose  knot, 
leaving  the  two  free  ends  which  are  to  be  attached  to  the 
myocardiograph  (Fig.  7). 

Place  the  board  with  the  turtle  on  the  stand  of  the  myo- 
cardiograph and  arrange  the  instrument  so  that  the  ends  of 
the  two  levers  (A  and  B)  shall  be  beside  the  ventricle.  Tie 
the  threads  attached  to  the  ventricle  to  the  ends  of  the  two 
levers.  The  writing  lever  now  begins  to  move  downwards 
with  each  contraction  and  upwards  with  each  relaxation  of 
the  ventricle.  Vary  its  height  so  that  it  will  record  the 
movements  of  the  heart  on  the  smoked  surface  of  a  drum 
properly  placed.  Arrange  the  stationary  lever  (D)  to  draw 
a  base  line  just  below  the  tracing  of  the  cardiac  movements. 

10-p 


144 


LABORATORY  GUIDE  IN 


After  taking  a  short  tracing  showing  the  normal  con- 
tractions of  the  heart,  take  one  showing  the  effects  of  stimu- 
lation of  the  vagus  with  both  weak  and  strong  currents. 

NOTE. — Mark   on  the   tracing  when   any  stimulation   is  begun  or  stopped  or 
when  any  drug  is  applied. 

Continuing  the  tracings,  apply  a  few  drops  of  a  pilocar- 


FIG.  No.    7.      Myocardiograph  for  turtle's  heart. 

pine  solution  (0.5  %}  to  the  heart.  When  the  contractions 
become  very  infrequent  or  stop  entirely,  apply  atropine 
(o.i  %)  to  the  heart,  adding  a  drop  or  two  at  a  time  to  the 
organ. 

When  the  contractions  return  to  normal,  stimulate  the 
vagus  with  the  electric  current  as  before. 

Apply  pilocarpine  again  to  the  heart.  Compare  the  trac- 
ings with  those  obtained  earlier  in  the  experiment. 


EXPERIMENTAL  PHARMACOLOGY.  145 


146  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  147 

Shake  3  or  4  cc.  of  chloroform  in  20  or  30  cc.  of  physio- 
logical salt  solution  and  pour  some  of  this  solution  from  time 
to  time  over  the  heart.  When  the  cardiac  contractions  be- 
come very  weak,  remove  the  chloroform  solution  with  a  piece 
of  absorbent  cotton  and  assist  the  recovery  of  the  heart  by 
bathing  it  in  fresh  salt  solution. 

Note  that  pilocarpine  does  not  stop  the  heart  after  atro- 
pine,  nor  has  vagus  stimulation  any  effect.  Chloroform,  on 
the  other  hand,  acts  after  atropine.  How  is  this  to  be  ex- 
plained ? 

Remove  carefully  the  paper  with  the  tracings  from  the 
drum  and  fix  it  by  passing  it  through  a  solution  of  shellac 
in  alcohol. 

Study  the  tracings  according  to  directions  given  on 
page  1 60. 

The  Effect  of  Drugs  on  Blood  Pressure. 

Anaesthetize  a  rabbit  or  a  cat  according  to  the  methods 
described  earlier  (page  n),  and  when  the  animal  is  com- 
pletely under  the  influence  of  the  anaesthetic,  tie  it  on  its 
back  on  the  operating  board  and  remove  the  hair  from  the 
neck  region.  Make  an  incision  about  four  inches  long  in 
the  median  line  of  the  neck  and  expose  the  trachea }  insert  a 
tube  (page  23)  to  allow  of  artificial  respiration  should  it 
become  necessary. 

Expose  the  jugular  vein  on  one  side  and  insert  a  cannula 
(page  19).  On,  the  opposite  side  expose  the  carotid  artery 
and  in  like  manner  tie  a  cannula  in  it  (page  20). 

Fill  the  venous  cannula  with  physiological  salt  solution 
and  the  carotid  cannula  with  a  solution  made"  by  mixing 
equal  volumes  of  water  and  a  saturated  solution  of  sodium 
sulphate. 


148  LABORATORY   GUIDE   IN 

The  ordinary  form  of  mercury  manometer,  which  is  so  well 
known  as  to  need  no  description,  is  used.  Fill  the  proximal 
limb  of  the  U-shaped  tube,  i.  e.,  the  limb  which  is  to  be  con- 
nected with  the  blood  vessel,  with  the  sodium  sulphate  solu- 
tion, using  for  this  purpose  a  pipette  with  a  long  tapering 
point.  The  lead  (or  rubber)  tube,  which  is  connected  to  the 
side  tube  of  the  proximal  limb  of  the  manometer,  is  to  be 
completely  filled  with  the  sulphate  solution,  taking  great 
care  to  see  that  all  air  is  expelled  from  the  tube  and  manom- 
eter. Close  the  clamp  on  the  tubing  which  connects  the 
lead  tube  and  manometer.  With  a  short  piece  of  rubber  tub- 
ing connect  a  large  pipette  (25  cc.)  filled  with  the  sulphate 
solution  to  the  upper  opening  of  the  proximal  limb  of  the 
manometer  and  blow  in  the  pipette  so  as  to  raise  the  pres- 
sure until  there  is  a  difference  of  10  or  12  cm.  in  the  height 
of  the  mercury  in  the  two  limbs.  Retain  this  pressure  by 
closing  the  piece  of  rubber  tubing  with  a  spring  or  screw 
clamp  and  disconnect  the  pipette. 

By  means  of  a  short  piece  of  rubber  tubing  connect  the 
distal  end  of  the  lead  tube  to  the  carotid  cannula  and  open 
the  clamps  so  as  to  allow  the  carotid  pulsations  to  be  trans- 
mitted to  the  mercury.  Arrange  the  writing  lever  so  that  it 
will  record  the  pulsations  on  a  blackened  drum  with  a  time 
marker  writing  below  the  tracing.  Keep  the  writing  lever 
in  light  contact  with  the  drum  by  means  of  a  small  weight 
suspended  by  a  thread  from  a  support  above  the  drum,  tak- 
ing care  that  the  thread  does  not  press  too  hard  against  the 
lever. 

Administer  the  following  drugs  in  the  manner  described, 
taking  a  short  piece  of  normal  tracing  before  each  is  given, 
and  waiting  until  the  effects  of  one  drug  have  passed  off 


EXPERIMENTAL  PHARMACOLOGY.  149 


LABORATORY   GUIDE   IN 


g-^gc,-^   m 


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f 

'%  /yj^  t^st^e^t  <#  <&~^c~*-^t~^c*^t^ 


EXPERIMENTAL  PHARMACOLOGY.  151 

before  applying  the  next.     Observe  the  precautions  as  to  the 
injection  of  any  air. 

1.  Amyl  nitrite:  3  drops  on  cotton  wool  applied  to  the 
animal's  nose  or  to  the  tracheal  tube  if  that  has  been  in- 
serted. 

2.  Chloroform :  given  on  cotton  wool  as  above. 

3.  Ether :  given  on  cotton  wool  as  above. 

4.  Suprarenal  extract.     Adrenalin  chloride   (i-iooo)  ;  I 
drop  in  salt  solution  injected  into  the  vein.     Wash  the  can- 
nula  and  syringe  thoroughly  before  injecting  the  next  drug. 

5.  Digitalis.     I  cc.  of  the  tincture  from"  which  the  alcohol 
has  been  evaporated  (given  intravenously). 

Repeat  this  injection  as  many  times  as  necessary,  noting 
the  successive  effects  on  the  tracing  until  the  heart  ceases. 

Allow  the  drum  to  continue  rotating  after  the  heart  has 
stopped,  so  that  the  writing  lever  traces  a  straight  line  from 
which  measurements  may  be  made  to  ascertain  the  absolute 
blood  pressure.  Remove  the  tracings  from  the  drums  and 
fix  them  by  passing  them  through  a  solution  of  shellac  in 
alcohol. 

The  blood  pressure  tracings  are  to  be  analyzed  according 
to  the  directions  given  on  page  160. 

Effect  of  Drugs  on  the  Dog's  Heart. 

Anaesthetize  a  dog  (page  12)  and  when  anaesthesia  is 
complete,  tie  the  animal  on  its  back  on  the  operating  table. 
Make  an  incision  through  the  skin  in  the  median  line  of  the 
neck,  and  after  exposing  the  trachea  insert  a  cannula  in  the 
usual  way  (page  23).  Expose  a  vein,  either  one  of  the  jug- 
ulars or  a  saphenous  in  the  leg,  and  insert  a  venous  cannula. 

Dissect  out  the  vagus  nerve  on  one  side,  tie  a  ligature 


152  LABORATORY  GUIDE   IN 

around  it  and  cut  the  nerve  between  the  ligature  and  the 
head 

Continue  the  skin  incision  in  the  median  line  from  the 
neck  down  the  thorax  to  an  inch  or  two  below  the  end  of  the 
sternum,  and  deepen  the  cut  to  the  bone.  Stop  all  bleeding 
points  with  artery  forceps. 

Connect  the  tubing  of  the  bellows  with  the  tracheal  tube 
and  start  artificial  respiration. 

Saw  through  the  entire  length  of  the  sternum,  keeping  ex- 
actly in  the  median  line  and  complete  the  division  with  bone 
forceps;  draw  the  two  sides  of  the  thorax  well  apart  and 
secure  them  to  the  sides  of  the  table  with  sharp  hooks. 

Regulate  the  amount  of  air  entering  the  lungs  by  means 
of  a  screw  clamp  placed  on  a  piece  of  rubber  tubing  on  the 
arm  of  the  tracheal  cannula. 

NOTE. — During  the  process  of  sawing  through  the  sternum  and  opening  the 
thorax  the  animal  must  be  very  deeply  under  the  anaesthetic,  otherwise  the 
heart  is  likely  to  go  into  delirium. 

Cut  both  phrenic  nerves  as  they  pass  down  to  the  di- 
aphragm lying  in  plain  sight  on  either  side  of  the  pericardial 
sac. 

Open  the  pericardium  along  its  entire  length  with  a  pair  of 
scissors  and  fasten  it  to  the  cut  edges  of  the  sternum  by 
means  of  four  or  five  stitches. 

.  Take  a  stitch  in  either  side  of  the  right  ventricle  with 
small  curved  needles  and  thread,  and  tie  the  thread  into  the 
heart  in  the  same  way  as  you  did  with  the  turtle.  Similarly 
take  stitches  in  the  right  auricle.  If  bleeding  should  occur 
from  the  needle  passing  completely  through  the  wall  of 
either  chamber  take  up  the  bleeding  point  with  a  pair  of  dis- 
secting forceps  and  tie  a  ligature  around  it. 


EXPERIMENTAL  PHARMACOLOGY. 


153 


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LABORATORY   GUIDE  IN 


£V^- 

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//r^'^~^L' 


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EXPERIMENTAL  PHARMACOLOGY. 


155 


Attach  the  two  sets  of  levers  of  the  dog's  myocardiograph1 
(Fig.  8)  to  the  heart  by  means  of  the  threads  and  arrange 
the  writing  levers  to  record  the  cardiac  movements  on  a 


FIG.  No.    8.      Myocardiograph  for  dog's  heart. 


1The  dog  myocardiograph  (Fig.  8)  is  constructed  on  exactly  the  same  prin- 
ciples as  is  the  myocardiograph  for  the  turtle's  heart  (Fig.  7),  excepting 
that,  as  is  shown  in  the  figure,  it  is  double,  so  that  it  will  record  the  move- 
ments of  both  auricle  and  ventricle.  The  apparatus  has  other  modifications  as 
is  seen  in  the  plate,  but  the  principle  is  not  essentially  altered.  A  descrip- 
tion of  that  part  of  the  apparatus  which  is  connected  to  the  heart  is  found  in 
the  Jour,  of  Physiol.,  1897,  XXI,  p.  213. 


156 


LABORATORY   GUIDE  IN 


blackened  drum.    Arrange  a  time  marker  to  write  just  below 
the  levers. 

Start  the  kymograph  and  take  a  normal  tracing,  after 
which  take  tracings  to  show  the  effects  of  the  following: 


FIG.  No.  9.      Anaesthetic  inhaler  for  dogs.      By   rotating  upper   part    of    apparatus 

"a"  slightly  upon  "b"  either  air  or  anaesthetic  may  be  forced  into  the 

animal's  lungs. 

1.  Vagus  stimulation,  weak  and  strong  currents. 

2.  Chloroform    (given    with    special   inhaler,    Fig.   9) ; 
continue  the  tracing  so  as  to  show  the  recovery. 

3.  Ether   (given  with  special  inhaler,  Fig.  9)  ;  continue 
the  tracing  so  as  to  show  the  recovery. 


EXPERIMENTAL  PHARMACOLOGY.  157 


158  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  159 

4.  Suprarenal  extract.      (Adrenalin  chloride,  i-iooo,  3 
drops  added  to  4  cc.  salt  solution  and  injected  into  the  vein 
with  a  syringe.) 

5.  Alcohol,  10  cc.  of  50  %  given  intravenously. 

6.  Digitalis,  tincture  evaporated  to  one-half  its  volume 
and  i  cc.  injected. 

7.  Caffeine,  0.2  G.  dissolved  in  hot  salt  solution. 

8.  Digitalis  (toxic  doses),  inject  the  concentrated  tinc- 
ture (see  6)  in  2  cc.  doses. 

The  heart  finally  becomes  irregular  and  may  pass  into  de- 
lirium. Mark  the  tracings  carefully  so  that  they  can  all  be 
identified  and  fix  them  in  shellac  as  before.  Analyze  them 
according  to  directions  given  on  page  163. 

Cut  off  the  head  of  the  animal  and  as  soon  as  convenient 
dissect  out  the  submaxillary  ducts  according  to  the  directions 
given  on  page  191.  This  dissection  will  allow  more  speedy 
operating  on  the  living  animal  in  the  experiment  on  the  sali- 
vary secretion. 

Remove  both  kidneys  leaving  as  much  of  their  vessels 
attached  as  possible  and  put  them  in  salt  solution  for  use  in 
Exp.,  page  167.. 
11-p 


160      .  LABORATORY   GUIDE  IN 


Analysis  of  Tracings. 

Graphic  records  are  obtained  in  three  experiments  (turtle's 
heart,  dog's  heart  and  blood  pressure)  and  it  is  of  first  im- 
portance that  these  records  should  be  carefully  analyzed  in 
order  to  properly  study  the  changes  induced  by  the  various 
drugs.  A  few  hints  are  given  below  to  call  attention  to 
some  of  the  more  important  points  to  be  looked  for,  and 
some  suggestions  are  made  as  to  the  best  way  of  studying 
the  tracings. 

Tracings  Obtained  from  the  Turtle's  Heart. 

It  is  rarely  necessary  in  these  records  to  take  accurate 
measurements,  as  the  changes  induced  by  the  drugs  are  gen- 
erally quite  marked. 

Describe  the  effect  of  stimulation  of  the  vagus,  noting 
whether  the  heart  stops  in  systole  or  diastole.  Likewise, 
describe  the  condition  of  the  heart  under  pilocarpine  (before 
and  after  atropine  administration)  and  also  the  effect  of  the 
atropine. 

Under  chloroform  note  especially  the  strength  of  the  heart 
as  indicated  by  the  extent  of  contraction  and  of  dilatation, 
which  is  measured  by  the  distance  of  the  tracing  from  the 
base  line. 

Summarize  the  actions  of  the  different  drugs. 

Tracings  from  the  Blood  Pressure  Experiment. 

Study  these  first  with  regard  to  changes  in  pressure.    Take 

careful  measurements  of  the  distance  of  the  tracing  from  the 

base  line  at  various  points  along  the  course  of  the  curve; 

viz.,  before  the  drug  is  injected  to  get  the  normal;  after  the 


EXPHRIMBNTAL  PHARMACOLOGY.  161 


^^^-iz^J-^U? 


162  LABORATORY  GUIDE   IN 


/ 


EXPERIMENTAL  PHARMACOLOGY.  163 

action  of  the  drug  becomes  apparent;  and  later  at  short  in- 
tervals of  time  which  will  depend  upon  the  drug  used. 
Where  the  curve  is  fairly  uniform  readings  every  thirty  sec- 
onds are  frequent  enough. 

Measuring  in  this  way  from  the  base  line  will  only  give 
relative  changes  in  pressure,  which  is  all  that  is  necessary  in 
these  experiments.  To  get  the  absolute  pressure,  the  zero 
line  which  was  marked  on  the  tracing  would  have  to  be  used, 
measuring  its  distance  above  or  below  the  base  line  and  add- 
ing or  subtracting  this  distance  from  the  figures  obtained 
above.  This  result  must  then  be  doubled  as  there  are  two 
arms  of  mercury  in  the  manometer. 

The  changes  in  the  rate  of  the  heart  are  studied  in  the 
following  manner:  Draw  two  lines  parallel  to  each  other 
and  perpendicular  to  the  base  line  so  that  they  shall  intersect 
the  tracing.  The  distance  of  the  lines  from  each  other  is 
determined  by  the  rate  of  the  drum  as  indicated  by  the  time 
marker;  they  are  usually  drawn  to  include  between  them  a 
space  equal  to  five  or  ten  seconds.  Count  the  heart  beats  in 
the  tracing  included  between  the  two  lines  and  estimate  the 
rate  of  the  heart  per  minute.  These  estimations  are  made 
along  the  curve  at  the  same  points  as  the  pressure  changes 
are  measured. 

Also  describe  any  other  changes  in  the  curves,  such  as  ir- 
regularities of  the  heart  or  of  the  respiration,  etc. 

Summarize  the  effects  of  the  different  drugs  upon  the 
pressure  of  the  ^lood  and  upon  the  rate  of  the  heart. 

Tracing  from  the  Dog's  Heart. 

These  are  to  be  studied  and  analyzed  by  a  combination  of 
the  methods  outlined  above.  The  rate  of  the  heart  is  ascer- 
tained as  described  under  the  directions  for  studying  the 


164  LABORATORY  GUIDE  IN 

blood  pressure  tracings,  while  the  strength  of  the  organ  as 
indicated  by  the  extent  of  systole  and  diastole  is  measured 
from  the  base  line.  Describe  all  other  changes  in  the  trac- 
ings, and  in  your  notes  make  a  full  summary  of  the  actions 
of  the  different  drugs. 


EXPERIMENTAL  PHARMACOLOGY.  165 


166  LABORATORY  GUIDU   IN 


EXPERIMENTAL  PHARMACOLOGY.  167 


Perfusion  of  the  Kidney. 

The  kidneys  which  were  removed  from  the  dog  (page  159) 
are  to  be  employed  in  the  following  experiment.  They 
should  not  be  kept  any  longer  than  is  absolutely  necessary 
after  being  removed  from  the  animal,  certainly  not  more 
than  twenty-four  hours,  as  after  that  time  post  mortem 
changes  have  advanced  so  far  as  to  interfere  with  the  action 
of  drugs  on  the  vessel  walls. 

In  preparing  the  kidneys  for  the  experiment  it  is  usually 
best  not  to  remove  the  capsule,  as  in  so  doing  the  paren- 
chyma is  frequently  torn.  Remove  the  fat  and  connective 
tissue  from  around  the  vessels  and  insert  glass  cannulas  in 
both  the  artery  and  the  vein.  Fill  the  vessels  and  cannulas 
with  salt  solution.  Fasten  a  small  glass  reservoir  (say  250 
cc.)  about  1.5  meters  above  the  table  and  connect  a  long 
piece  of  rubber  tubing  with  its  lower  orifice. 

Strain  some  defibrinated  blood  through  a  cloth  and  dilute 
it  with  two  volumes  of  saline  solution.  (Instead  of  diluted 
blood,  Locke's  solution  may  be  used.) 

Put  about  loo  cc.  of  this  diluted  blood  in  the  reservoir  and 
allow  it  to  fill  the  rubber  tube  and  then  clamp  the  latter. 
Connect  the  lower  end  of  the  rubber  tube  with  the  arterial 
cannula  taking  care  that  all  air  is  expelled.  Remove  the 
clamp  from  the  tubing  and  place  a  receptacle  under  the 
venous  cannula  to  catch  the  fluid  after  it  has  passed  through 
the  kidney.  Measure  the  fluid  (blood)  in  this  receptacle  at 
intervals  of  five  minutes  during  the  course  of  the  experi- 
ment. After  a  fairly  constant  rate  has  been  determined  by 
two  or  three  readings,  put  into  the  blood  reservoir  O.I  G. 


168  LABORATORY  GUIDE  IN 

sodium  nitrite  (i  cc.  of  10  %  solution)  and  mix  the  solution. 
The  experiment  may  be  hastened  by  disconnecting  the  rub- 
ber tubing  from  the  cannula  and  allowing  the  normal  diluted 
blood  in  the  tube  to  escape  and  to  be  replaced  by  sodium 
nitrite  blood,  and  then  connecting  up  the  tube  and  cannula 
again. 

Take  several  readings  of  the  rate  of  the  outflow  of  the 
blood  under  the  nitrite  action.  When  a  constant  figure  has 
been  reached,  replace  the  nitrite  blood  in  the  reservoir  and 
rubber  tube  by  normal  diluted  blood  and  allow  it  to  flow 
through  the  kidney  to  wash  out  all  the  sodium  nitrite. 

Now  add  to  the  normal  blood  in  the  reservoir  some  ad- 
renalin chloride  (i  cc.  of  i-iooo  solution)  and  measure  the 
outflow  under  its  action. 

What  do  the  changes  in  the  rate  of  outflow  under  the  two 
drugs  indicate  in  regard  to  their  action  on  the  blood  vessels  ? 
How  do  the  results  help  to  explain  the  observations  made  in 
the  experiment  on  blood  pressure  (page  151). 

Diuresis. 

Anaesthetize  a  rabbit  with  paraldehyde  (page  n)  and 
when  anaesthesia  is  complete  tie  the  animal  on  the  operating 
board.  Cut  the  hair  from  the  neck  and  also  from  the  lower 
part  of  the  abdomen.  Expose  a  jugular  vein  and  insert  a 
cannula  as  usual. 

The  effect  of  drugs  on  the  excretion  of  the  urine  is  to  be 
studied  in  relation  to  the  drugs  used  and  also  with  regard  to 
the  height  of  the  blood  pressure.  In  order  to  do  this  expose 
a  carotid  artery  and  insert  a  cannula  into  it  and  connect  the 
cannula  with  the  mercury  manometer  which  has  been  ar- 


EXPERIMENTAL  PHARMACOLOGY.  169 

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170  LABORATORY  GUIDE   IN 

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EXPERIMENTAL   PHARMACOLOGY.  171 

ranged  according  to  the  directions  given  under  "Blood  Pres- 
sure," page  147. 

To  insert  a  cannula  in  the  bladder  to  collect  the  urine, 
make  an  incision  about  an  inch  and  a  half  long  in  the  median 
line  of  the  abdomen,  beginning  the  incision  at  the  symphysis 
and  extending  it  upward  the  distance  named.  Deepen  the 
cut  until  the  peritoneal  cavity  is  opened.  The  bladder  is 
usually  found  partially  distended  and  lying  amid  folds  of  fat. 
Draw  it  out  through  the  opening  made  and  empty  it'  by 
gentle  pressure,  catching  the  urine  in  a  dish  as  it  flows  from 
the  urethra. 

Tie  a  knot  loosely  in  a  piece  of  thread  and  lay  it  on  the 
abdomen  so  that  the  knotted  thread  will  encircle  the  bladder 
and  be  ready  to  tie  the  cannula  in  place  when  the  latter  shall 
have  been  inserted. 

After  locating  the  position  of  the  ureters,  so  as  to  avoid 
including  them  in  the  ligature,  take  hold  of  the  upper  sur- 
face of  the  bladder  on  the  two  sides  with  two  pairs  of  forceps 
and  draw  them  gently  up  and  outwards  so  as  to  exert  a 
slight  tension  on  the  organ.  While  one  operator  holds  the 
bladder  in  this  way  the  other  operator  makes  a  cut  with  a 
sharp  pair  of  scissors  midway  between  the  forceps  in  the 
fundus  of  the  bladder.  The  cut  will  vary  with  the  size  of 
the  cannula  flange  (a,  Fig.  10)  and  will  probably  average 
about  two  centimeters  in  length.  Into  this  incision  now 
place  the  cannula  so  that  its  flange  shall  be  entirely  sur- 
rounded by  bladder  tissue  and  tie  it  in  place  with  the  loose 
ligature  already  mentioned.  Examine  it  after  it  is  tied  to 
see  that  the  ureters  are  not  included. 

Fill  the  bladder  and  cannula  with  the  urine  which  was  ob- 
tained earlier  or  with  warm  physiological  salt  solution  and 


172 


LABORATORY  GUIDE  IN 


connect  the  end  of  the  cannula  with  the  piece  of  bent  tubing 
(b,  Fig.  10)  which  can  now  rest  on  a  support  so  as  to  ex- 
tend over  the  end  of  the  operating  board  where  small 
weighed  dishes  are  to  be  placed  to  catch  the  urine. 

As  soon  as  the  urine  begins  to  drop 
from  the  tube  a  record  of  the  time  is  to 
be  made,  and  thereafter  to  the  end  of 
the  experiment,  every  five  minutes,  the 
dish  with  the  urine  is  to  be  replaced 
by  a  clean  receptacle  and  the  first  one 
weighed,  and  the  weight  of  urine  re- 
corded. 

The  blood  pressure  tracing  is  to  be 
taken  on  a  very  slow  drum  upon  which 
the  five  minute  intervals  are  to  be 
marked  under  the  corresponding  trac- 
ing, and  the  amount  of  urine  for  that 
period  is  also  to  be  marked  within  the 
intervals  indicated.  By  keeping  the 
record  in  this  way  the  drum  will  show 
the  drug  used,  the  time,  the  amount  of 
urine  excreted  in  the  different  intervals 
and  the  height  of  the  blood  pressure 
during  the  same  periods. 
FIG.  No.  10.  Bladder  Take  two  or  three  readings  until  a 
CTulaM  ^'  Fla"get°bc  fair  regularity  is  obtained,  and  after 

tied  m  bladder.       B.    Bent  J 

delivery  tube.  this  normal  is  ascertained  inject  into  the 

vein  40  mg.  of  caffeine  dissolved  in  hot  water. 

After  this  injection  the  urine  probably  will  be  increased 
and  the  next  drug  must  not  be  injected  until  the  flow  has  re- 
turned to  about  the  normal. 


EXPERIMENTAL  PHARMACOLOGY.  173 


174  LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  175 

When  the  amount  of  urine  has  thus  fallen,  inject  0.250  G. 
(5  cc.  of  a  5%  solution)  of  sodium  nitrate  and  collect  the 
urine  as  before. 

When  normal  is  reached  once  more,  inject  0.250  G.  (5  cc. 
of  5  %  solution)  of  sodium  chloride  and  compare  the  results 
with  the  nitrate. 

NOTE. — If  time  allows,  the  diuretic  effect  of  digitalis  may  be  studied  in  the 
same  way. 

When  the  experiment  is  concluded  fix  the  tracings  and 
later  study  them,  not  only  as  regards  the  drug  injected,  but 
also  in  relation  to  the  changes  in  blood  pressure. 

Caffeine,  digitalis  and  sodium  nitrate  are  taken  as  repre- 
sentatives of  the  large  class  of  drugs  known  as  Diuretics 
from  their  action  in  increasing  the  amount  of  urine. 

They  may  act  either  on  the  heart  or  on  the  kidney,  and 
1;hey  are  known  as  cardiac  or  renal  diuretics  according  to  the 
point  of  action.  Digitalis  is  an  example  of  the  former  class, 
as  it  causes  diuresis  indirectly  through  its  effects  on  the  cir- 
culation. Caffeine  probably  acts  directly  on  the  renal  epi- 
thelium, while  various  salts,  of  which  sodium  nitrate  is  an 
example,  induce  diuresis  by  their  salt  action  (saline  diuretic) 
in  addition  to  which  there  may  be  a  stimulant  action  on  the 
kidney  epithelium. 

Therapeutic  uses  of  the  Diuretics. 

See  also  under  Caffeine  and  Digitalis,  Therapeutic  uses, 
(pages  100  and  128). 

These  drugs  or  their  allies  are  chiefly  employed  in  diseased 

conditions  in  which  the  amount  of  urine  excreted  is  below  the 

normal,  a  condition  which  may  result  from  disease  of  the 

kidneys  or  of  the  heart.   In  conditions  of  oedema  or  ascites, 

12-p 


176  LABORATORY   GUIDE  IN 

there  is  an  abnormal  collection  of  fluid  in  the  tissues ;  its 
removal  may  be  aided  by  the  use  of  the  diuretics.  In  poison- 
ing with  an  irritant  drug  which  is  eliminated  by  the  kidneys, 
the  latter  may  be  protected  from  the  action  of  the  toxic 
substance  by  increasing  the  volume  of  urine  through  the  ad- 
ministration of  these  drugs. 


EXPERIMENTAL  PHARMACOLOGY.  177 


LABORATORY   GUIDE   IN 


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v/ 


EXPERIMENTAL   PHARMACOLOGY.  179 


The  Belladonna  Series. 

Several  plants  belonging  to  the  Solanacese  furnish  alka- 
loids which  are  very  closely  related,  not  only  chemically,  but 
also  in  their  pharmacological  effects.  Chief  among  these 
alkaloids  is  Atropine  which  is  contained  in  the  leaves  and 
roots  of  the  deadly  nightshade  (Atropa  Belladonna).  Next 
in  importance  to  atropine  are  Hyoscyamine  and  Hyoscine. 
These  alkaloids  are  all  obtained  from  other  sources  than 
that  named,  viz.;  henbane  (Hyoscyamus  niger),  the  thorn- 
apple  (Datura  Stramonium)  and  other  plants  of  less  im- 
portance. They  are  very  hard  to  isolate  in  absolutely  pure 
form  because  hyoscyamine  is  very  prone  to  change  to  atro- 
pine during  the  manipulations. 

As  stated  above,  they  resemble  one  another  closely  in  their 
properties  and  actions,  so  that  atropine  alone  will  be  studied. 

Examine  Belladonna  leaves  and  roots. 

I.  Prepare  the  alcoholic  extract  of  belladonna  leaves  ac- 
cording to  the  U.  S.  P.,  using  50  G.  of  the  powdered  drug. 

Define  an  "extract"  of  a  drug. 

II.  Take  some  of  the  extract,  an  amount  about  the  size  of 
a  pea  and  rub  it  up  in  5  cc.  of  salt  solution  and  put  a  drop  of 
the  watery  extract  in  a  cat's  eye  and  watch  for  any  changes 
comparing  it  with  the  normal  eye.     The  cat  is  best  put  in  a 
cat  box  (footnote,  page  12)  with  the  head  exposed  during 
the  experiment;  drop  the  solution  near  the  outer  canthu?oi 
the  eye  so  that  it  will  have  to  cross  the  eyeball  before  escap- 

i  ing\by  the  nasal  duct. 

*This  action  of  the  atropine  series  upon  the  pupil  is  known 
as  mydriasis  and  the  drugs, producing  it  are  mydriatics.     It 


180  LABORATORY  GUIDE,  IN 

is  the  most  delicate  test  known  for  the  group,  although  a 
few  other  drugs,  of  which  cocaine  is  the  most  important,  act 
as  mydriatics,  though  to  a  less  marked  degree. 

III.  Isolate  the  atropine  from  the  extract  of  belladonna 
by  the  "shaking-out"  process  as  described  below.  Observe 
the  precautions  mentioned  under  the  isolation  .of  strychnine, 
(page  68). 

Mix  the  extract  thoroughly  with  about  100  cc.  of  distilled 
water  and  render  it  alkaline  with  ammonia.  Put  the  mix- 
ture in  a  separating  funnel  and  add  20  cc.  of  chloroform. 
Shake  gently  for  3  to  5  minutes  and  set  the  funnel  aside  to 
allow  the  chloroform  to  separate,  drawing  it  off  into  a  flask. 
Add  15  cc.  more  chloroform  to  the  watery  extract  and  shake 
as  before,  and  draw  off  the  chloroform  into  the  flask  with  the 
first  lot.  The  watery  extract  can  now  be  thrown  away. 

Put  the  chloroform  containing  the  alkaloid  in  the  separa- 
ting funnel  and  add  about  50  cc.  of  water  rendered  distinctly 
acid  with  sulphuric  acid.  Agitate  the  mixture  several  min- 
utes as  before,  drawing  off  the  chloroform  from  the  water 
when  separation  has  taken  place.  Repeat  the  process  with 
the  chloroform  and  50  cc.  of  fresh  acidulated  water.  (The 
waste  chloroform  is  to  be  placed  in  a  jar  on  the  side  table 
for  redistillation.)  If  the  watery  solution  is  colored,  shake 
it  in  a  flask  with  powdered  charcoal  and  filter  it.  The  fil- 
trate containing  the  atropine  is  now  rendered  alkaline  with 
ammonia  and  shaken  out  again  with  20  cc.  of  chloroform, 
which  is  drawn  off  into  an  evaporating  dish.  Repeat  the 
process  with  10  cc.  of  chloroform  and  add  it  to  the  20  cc.  in 
the  dish  and  set  the  whole  amount  aside,  protected  from  the 
dust  to  allow  the  chloroform  to  evaporate  and  the  atropine 
to  crystallize. 

IV.     Place  a  few  crystals  of  atropine   (from  Exp.  Ill) 


EXPERIMENTAL  PHARMACOLOGY. 


181 


182  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY. 


183 


on  a  porcelain  cover  and  add  to  them  a  drop  of  concentrated 
nitric  acid  and  evaporate  it  to  dryness  over  the  water  bath. 
Allow  the  cover  to  cool  and  then  touch  the  residue  with  a 
glass  rod  which  has  been  dipped  in  an  alcoholic  solution  of 
caustic  potash.  A  rich  violet  color  is  produced,  changing  to 
a  dark  red,  which  finally  fades  away,  but  can  be  reproduced 
by  the  addition  of  fresh  alcoholic  potash.  This  reaction 
(known  as  Vitali's  test)  is  almost  peculiar  to  the  atropine 
series  and  is  said  to  be  given  by  o.oooi  mg.  of  the  alkaloid. 

V.  Inject  into  a  frog  15  mg.  of  atropine  sulphate  and  de- 
scribe all  the  symptoms  which  are  observed  following  the  in- 
jection of  the  drug.     Keep  the  animal  for  several  days  until 
complete  recovery  takes  place.     Compare  the  effects  with 
those  induced  by  any  other  drugs  used  earlier  in  the  course. 

VI.  Dissolve  the  remainder  of  the  atropine  in  about  20 
cc.  of  distilled  water  which  has  been  rendered  acid  with  sul- 
phuric or  hydrocholoric  acid.     Divide  the  solution  into  five 
parts,  put  it  in  test  tubes  and  add  the  alkaloidal  precip- 
itants  as  follows: 


Reagent. 

Precipitate? 

a 

Tannic  acid. 



b 

Picric  acid. 

^~ 

c 

Iodine  in  potassium  iodide. 

+ 

d 

Mercury-potassium  iodide. 

•/- 

e 

Phosphotungstic  acid. 

-/- 

VII.  Take  o.ooi  G.  of  atropine  sulphate  (which  will  be 
furnished  you)  and  describe  all  the  symptoms  experienced. 
Count  the  rate  of  your  heart  before  and  after  taking  the 
drug  and  see  if  there  is  any  change. 

The  action  of  atropine  on  the  innervation  of  the  heart  was 
studied  on  the  turtle  (page  144)  ;  and  on  the  rabbit  (page 
139)- 


184  LABORATORY   GUIDE  IN 

Its  action  on  secretions  will  be  studied  on  the  salivary 
gland  (page  196). 

Its  mydriatic  effect  when  applied  locally  was  shown  in 
Exp.  II,  and  the  same  effect  from  internal  administration 
is  studied  under  the  heading  "Cervical  Sympathetic  Nerves," 
(page  188). 

Examine  Atropa  Belladonna, 
Hyoscyamus  niger, 
Datura  Stramonium, 
the  alcoholic  extract  of  belladonna  leaves, 
tincture  of  belladonna  leaves  and 
the  extract  and  tincture  of  hyoscyamus. 

Therapeutic  Uses. 

Owing  to  the  fact  that  atropine  has  such  a  wide  range  of 
activities,  its  uses  in  medicine  are  very  numerous.  Among 
these  a  few  may  be  mentioned  as  being  indicated  by  the  va- 
rious experiments  in  which  the  drug  is  employed.  It  is  used 
as  a  stimulant  in  the  depressed  state  of  the  central  nervous 
system  occufring  in  shock,  collapse  and  in  narcotic  poison- 
ing. (Compare  strychnine  and  caffeine.)  For  its  peri- 
pheral action  it  is  employed  for  many  purposes.  In  cases  of 
extreme  slowness  of  the  heart  ( Brady cardia)  due  to  over- 
activity  of  the  vagus,  atropine  may  be  employed.  It  is  used 
to  dilate  the  pupil  of  the  eye  to  allow  of  ophthalmoscopic 
examination,  or  as  a  therapeutic  agent  in  the  treatment  of 
many  eye  diseases. 

In  excessive  secretion  of  the  saliva  such  as  occurs  in 
poisoning  with  mercury,  and  in  the  night  sweats  of  tuber- 
culosis, atropine  is  very  valuable. 

Finally  it  is  used  in  cases  of  poisoning  by  pilocarpine  or 
muscarine,  the  latter  being  an  alkaloid  resembling  pilocar- 
pine in  many  of  its  actions. 


EXPERIMENTAL  PHARMACOLOGY.  185 


LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  187 


Action  of  Drugs  on  the  Cervical  Sympathetic  Nerves. 

Anaesthetize  a  rabbit  (an  albino  if  possible)  with  paralde- 
hyde  (page  n)  and  when  anaesthesia  is  complete  tie  the 
animal  on  the  operating  board  and  remove  all  the  hair  from 
the  neck  region.  Expose  the  jugular  vein  on  one  side  and  in- 
sert a  cannula.  Expose  the  carotid  sheath  on  the  other  side, 
open  it,  laying  bare  the  vessels  and  nerves.  The  largest 
nerve  seen  is  the  vagus  and  the  smallest  is  the  depressor 
nerve,  while  the  third  is  the  cervical  sympathetic  trunk,  which 
is  to  be  separated  very  carefully  from  all  connective  tissues 
and  a  fine  thread  passed  under  it.  Make  sure  you  have 
the  correct  nerve  by  gently  raising  it  and  stimulating  it  with 
a  weak  tetanizing  current,  observing  meantime  the  effect  on 
the  pupil  on  the  corresponding  side ;  it  should  dilate  during 
the  stimulation.  If  the  correct  nerve  has  been  isolated  tie 
the  ligature  around  it  as  low  down  in  the  neck  as  convenient 
and  cut  the  nerve  trunk  below  the  ligature.  Gently  hold  the 
animal's  ears  up  against  the  light  so  that  the  vessels  may  be 
clearly  seen,  and  observe  the  difference  in  their  size  on  the 
two  sides ;  note  also  the  difference  in  the  temperature  of  the 
two  ears.  Compare  the  size  of  the  pupils  on  the  two  sides. 
How  would  you  explain  these  changes? 

Raise  the  trunk  of  the  cut  nerve  carefully  by  the  ligature 
and  stimulate  it  with  the  electric  current  as  before.  During 
this  stimulation  look  especially  for  any  changes  in  the  calibre 
of  the  ear  vessels  on  that  side;  and  for  any  changes  in  the 
position  of  the  eyeball,  or  of  the  eyelids,  or  in  the  size  of  the 
pupil.  After  the  changes  have  been  seen  and  described,  in- 
ject into  the  jugular  vein  3  mg.  of  nicotine  chloride.  Im- 


188  LABORATORY  GUIDE  IN 

mediately  after  the  injection  notice  the  effect  of  the  drug  on 
the  animal's  respiration,  and  also  look  for  any  movement  in 
the  animal's  whiskers. 

Again  stimulate  the  cut  cervical  sympathetic  comparing 
the  results  with  those  obtained  at  the  first  stimulation. 
(Should  they  be  the  same  as  before,  it  will  be  necessary  to 
inject  more  nicotine.)  Now  using  very  careful  dissection 
trace  the  trunk  of  the  sympathetic  toward  the  head  until  the 
superior  cervical  ganglion  is  reached.  It  will  be  recognized 
as  a  small  whitish  enlargement  on  the  trunk  of  the  nerve. 
When  it  is  found,  stimulate  the  trunk  of  the  sympathetic  be- 
yond the  ganglion,  that  is,  between  the  ganglion  and  the 
head  of  the  animal.  Compare  the  results  of  stimulation  at 
this  point  with  those  obtained  below  the  ganglion. 

NOTE. — In  addition  to  the  nerve  fibres  which  run  to  the  eye  by  way  of  the 
cervical  sympathetic  and  which  are  distributed  to  the  radial  muscle  fibres  of 
the  iris  there  are  nerves  which  control  the  circular  fibres.  These  come  from 
the  motor  oculi  to  the  ciliary  ganglia  and .  run  from  that  point  to  the  iris. 
The  size  of  the  pupil  can  be  changed  by  drugs  acting  along  the  course  of 
these  fibres  also. 

Inject  into  the  jugular  vein  5  mg.  of  pilocarpine  (i  cc.  of 
0.5%  solution)  and  observe  any  change  in  the  pupil.  (Note 
any  effect  on  the  heart  rate.)  Finally  inject  I  mg.  atropine 
sulphate  (i  cc.  of  0.1%  solution).  What  effect  has  it  upon 
the  pupil  ? 

What  effect  has  nicotine  upon  the  sympathetic  ganglia? 

Cocaine  dilates  the  pupil  by  stimulating  the  terminations 
of  the  nerves  from  the  superior  cervical  ganglia  which  end 
on  the  radial  fibres. 

Physostigmine  (Eserine)  acts  on  the  same  point  and  in 
the  same  manner  as  pilocarpine,  i.  e.,  on  the  nerve  termina- 
tions on  the  circular  muscular  fibres;  atropine  acts  at  the 
same  place,  but  instead  of  stimulating  the  nerve  endings,  its 
action  is  to  paralyze  them,  removing  the  pilocarpine  effects. 


EXPERIMENTAL  PHARMACOLOGY.  189 


*£-  ' 


190 


LABORATORY  GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  191 

Therapeutic  Uses. 

Physostigmine  (eserine)  and,  less  commonly,  pilocarpine 
are  used  in  ophthalmological  practice  to  contract  the  pupil  of 
the  eye  in  certain  diseased  conditions,  the  most  important  of 
which  is  glaucoma.  For  this  purpose  a  solution  of  l/\  to  I  % 
is  dropped  in  the  eye  as  often  as  necessary. 

Atropine  is  used  to  dilate  the  pupil  to  permit  of  ophthal- 
moscopic  examinations,  and  as  a  therapeutic  agent  in  many 
ocular  diseases.  Homatropine,  an  artificial  alkaloid  re- 
sembling atropine  in  its  action  on  the  eye,  is  largely  used  as  a 
substitute  for  the  latter,  as  its  action  is  not  nearly  as  pro- 
longed. 

Cocaine  is  also  largely  employed  to  dilate  the  pupil  to 
allow  of  examination  of  the  interior  of  the  eye. 

Mydriatics  are  drugs  which  dilate  the  pupil ;  myotics  con- 
tract it. 

Salivary  Secretion. 

Anaesthetize  a  large  dog  with  morphine  and  chloretone 
(page  12)  and  if  necessary  supplement  them  by  the  use  of 
chloroform.  When  anaesthesia  is  complete,  expose  a  vein 
in  the  leg  and  insert  a  cannula.  As  a  matter  of  precaution, 
the  trachea  may  also  be  exposed  and  a  cannula  tied  in  it  to 
permit  of  artificial  respiration  should  it  become  necessary. 

The  submaxillary  duct  and  chorda  tympani  nerve  are  to  be 
dissected  out  according  to  the  following  directions.  (This 
dissection  should  have  been  previously  carried  out  on  the 
head  of  a  dead  animal  (from  Exp.,  page  159)  so  as  to  allow 
greater  speed  in  operating  on  the  living  dog). 

"Make  an  incision  3  or  4  inches  long  through  the  skin  and 
platysma  muscle  along  the  inner  border  of  the  lower  jaw,  be- 
13-p 


192 


LABORATORY   GUIDE  IN 


ginning  about  the  angle  of  the  mouth  and  continuing  back- 
ward towards  the  angle  of  the  jaw.  Legate  doubly  and 
divide  any  vessels  that  come  in  your  way.  Divide  the  digas- 
tric muscle  about  its  anterior  third  and  clear  it  from  its  at- 
tachment. The  broad,  thin  mylo-hyoid  will  now  be  seen 
with  its  motor  nerve  lying  on  it.  Divide  the  muscle  across 


FIG.  No.  ii.      Dog's  head  dissected  to  show  Chorda  Tympani.      A.   Chorda  Tym- 

pani,  B.  Wharton's  duct,  C.    Hypoglossal   nerve,    D.    Mylo-hyoid   muscle 

turned  back,  E.  Lingual  nerve,  F.  Digastric  muscle  cut,  G.  Ra- 

mus  of  jaw  cut  away  to   show  nerve   emerging  from 

behind  it. 

its  middle  and  dissect  it  up  carefully.  The  lingual  nerve  is 
seen  emerging  from  under  the  ramus  of  the  jaw,  running 
transversely  towards  the  middle  line  and  then  passing  for- 
ward parallel  to  the  larger  hypoglossal  nerve.  In  its  trans- 
verse course  it  crosses  the  submaxillary  and  sublingual  ducts. 


EXPERIMENTAL  PHARMACOLOGY. 


193 


194  LABORATORY   GUIDE  IN 


EXPERIMENTAL   PHARMACOLOGY.  195 

The  chorda  tympani  leaves  the  lingual  nerve  and  runs  back- 
ward along  the  duct  towards  the  gland.  Put  a  glass  cannula 
into  the  duct  in  the  same  way  as  you  would  put  a  cannula  in 
a  vein.  Trace  back  the  lingual  and  put  a  ligature  around  it 
and  tie  it  as  far  back  as  possible,  and  then  cut  it  centrally  so 
as  to  permit  of  the  chorda  tympani  being  put  on  electrodes." 
(Stewart.)  Connect  the  cannula  with  a  long,  horizontal 
tube  on  which  a  scale  is  marked. 

While  the  dissection  of  the  duct  is  being  carried  on,  a 
second  operator  is  to  dissect  out  the  cervical  sympathetic 
trunk  on  the  same  side.  In  the  dog  the  cervical  sympathetic 
is  contained  in  the  carotid  sheath  and  closely  connected  with 
the  vagus,  from  which  it  may  be  separated  by  careful  dis- 
section, as  the  fibres  run  in  distinct  strands.  For  some  dis- 
tance below  the  superior  cervical  ganglion  the  two  nerve 
trunks  are  not  connected.  After  the  sympathetic  is  isolated 
tie  a  ligature  around  it  and  cut  the  nerve  below  it. 

If  the  experiment  has  been  properly  done,  both  nerves 
(viz.,  the  chorda  and  the  sympathetic)  can  be  stimulated  by 
placing  them  on  electrodes,  and  any  secretion  from  the  gland 
will  pass  into  the  horizontal  tube,  where  it  can  be  measured 
on  the  scale. 

1.  Stimulate  the  chorda  tympani  with  a  weak  induction 
current  and  watch  the  effect  on  the  secretion. 

2.  When  the  effect  of  chorda  stimulation  has  passed  off 
stimulate  the  sympathetic  trunk  (central  or  head  end)  and 
observe  the  difference  in  the  quantity  and  quality  of  the 
saliva  when  compared  with  the  "chorda  saliva." 

3.  'Inject  30  mg.  of  nicotine  chloride  and  observe  closely 
the  effect  on  the  secretion. 

4.  Stimulate  the  chorda  again  and  compare  the  results 
with  those  obtained  in  i. 


196  LABORATORY   GUIDE   IN 

5.  Stimulate  the  sympathetic  and  compare  with  2. 

6.  Trace  the  trunk  of  the  sympathetic  to  the  superior 
cervical  ganglion  and  stimulate  the  nerve  beyond,  observing 
if  secretion  can  be  obtained. 

Inject  the  following  drugs  in  the  order  named,  observing 
their  effects  on  the  secretion: 

7.  Nicotine,  5  mg.,  compare^with  3. 

8.  Pilocarpine,  5  mg. 

9.  Atropine,  i  mg. 

10.  Pilocarpine,  5  mg.,  compare  with  8. 

11.  Suprarenal  extract,  Adrenalin  chloride,   i-iooo  sol., 
5  drops. 

What  effect  has  nicotine  upon  the  sympathetic  ganglia? 
Compare  also  the  experiment  on  the  sympathetic  ganglia  in 
the  rabbit  (page  187). 

As  was  shown  in  this  experiment,  the  salivary  gland  is 
innervated  from  two  sources,  both  of  which  are  acted  on  by 
nicotine.  Pilocarpine  and  atropine  affect  the  salivary  secre- 
tion by  acting  on  the  cranial  nerve  supply  of  the  gland.  The 
action  of  the  suprarenal  gland  extract  in  causing  secretion  is 
not  as  well  known,  but  it  is  believed  to  be  upon  the  termina- 
tions of  the  sympathetic  fibres  in  the  gland  cells.  (It  is  not 
always  possible  to  get  secretion  of  saliva  by  the  injection  of 
suprarenal  extract.) 


EXPERIMENTAL  PHARMACOLOGY.  197 


198  LABORATORY  GUIDE  IN 


HXPZRIMZNTAL  PHARMACOLOGY.  199 


The  Sphygmomanometer. 

Among  the  pharmacological  experiments  which  the  stu- 
dent may  carry  out  upon  himself,  there  are  very  few  which 
allow  of  absolutely  accurate  records  being  kept  such  as  are 
obtained  in  animal  experimental  work.  The  recent  develop- 
ment of  instruments  by  which  the  human  blood  pressure  may 
be  determined  opens  up  a  new  field  in  which  the  student 
may  study  upon  himself  the  effects  of  drugs  whose  actions 
he  has  demonstrated  upon  the  lower  animals. 

Any  of  the  newer  sphygmomanometers,  such  as  those  de- 
vised by  Brlanger,  Stanton,  Janeway  or  the  Cook  modifica- 
tion of  the  Riva-Rocci,  may  be  employed  for  this  purpose. 
The  underlying  principle  is  essentially  the  same  in  all,  and 
for  complete  descriptions  of  these  instruments  reference 
must  be  made  to  special  works  on  the  subject.1 

The  upper  arm  of  the  "patient"  is  encircled  by  a  broad 
rubber  bag  (12-15  cm-  wide)  connected  by  rubber  tubing 
with  a  mercury  manometer  and  a  rubber  bulb.  The  pressure 
in  the  closed  system  of  tubing  is  raised  by  means  of  the  bulb 
until  the  pressure  in  the  bag  is  sufficient  to  obliterate  the 
radial  pulse  under  the  finger  of  the  operator.  The  pressure 
is  now  lowered  slowly  until  the  pulse  reappears  and  then 
the  pressure  may  be  raised  to  cause  the  pulse  to  disappear 
once  more.  In  this  way  the  systolic  or  maximum  pressure 
may  be  obtained  and  recorded  by  observing  the  height  of  the 
mercury  column. 

With  some  of  the  instruments  the  minimum  or  diastolic 


JThe  Clinical  Estimation  of  Blood  Pressure.     Janeway,  New  York,  1904. 


200  LABORATORY  GUIDE  IN 

pressure  may  also  be  estimated,  but  this  hardly  necessary  in 
the  following  experiments. 

A  few  precautions  are  necessary  and  they  may  be  given 
most  concisely  by  saying  that  the  subject  of  the  experiment 
should  as  far  as  possible  refrain  from  all  mental  or  physical 
exertion.  During  the  course  of  the  experiment  he  should 
sit  or  lie  quietly  and  take  no  interest  whatever  in  his  sur- 
roundings. It  is  usually  most  satisfactory  to  have  three 
students  work  together,  one  to  receive  the  drug,  one  to  man- 
age the  air  bulb  and  take  the  readings,  and  the  third  to  direct 
his  attention  entirely  to  the  pulse. 

The  frequency  of  the  readings  and  the  length  of  time  over 
which  they  should  be  taken  will  depend  upon  the  drug  used. 
For  instance,  the  effects  of  amyl  nitrite  may  be  said  to  be 
over  in  a  very  few  minutes,  but  during  that  time  the  readings 
should  be  taken  as  frequently  as  possible.  With  the  other 
drugs  used,  readings  should  be  made  every  five  minutes  and 
if  possible  they  should  be  continued  for  an  hour  or  two  or 
until  the  pressure  has  returned  to  the  normal  height  again. 
In  any  case,  two  or  three  estimations  of  the  normal  pressure 
should  be  made  before  the  drug  is  administered. 

The  experiments  mentioned  below  may  be  varied  almost 
indefinitely  by  using  the  drugs  in  varying  dosage  or  in  com- 
bining two  drugs  in  one  experiment,  administering  the  sec- 
ond after  the  effect  of  the  first  is  plainly  manifest. 

The  pulse  rate  is  to  be  counted  and  recorded  together 
with  the  pressure  readings  on  the  blanks  provided. 

The  following  drugs  are  suggested : — 

Amyl  nitrite,  I  to  3  drops  given  by  inhalation. 

Strychnine  sulphate,  0.002  G.  to  0.005  G.,  by  hypodermic 
or  by  mouth. 


UXPBRIMHNTAL   PHARMACOLOGY.  201 


202  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  203 

Atropine  sulphate,  0.0005  G.  to  o.ooi  G.,  by  hypodermic 
or  by  mouth. 

Sodium  nitrite,  o.i  G.  by  mouth. 

Nitroglycerin,  I  to  3  drops  by  mouth. 

Digitalin,  o.ooi  G.,  by  mouth  or  hypodermic. 

The  effects  of  smoking  upon  the  blood  pressure  may  be 
tested  upon  a  suitable  individual;  the  most  pronounced  re- 
sults will  probably  be  obtained  from  inhaling  the  smoke  from 
a  "strong"  pipe  or  cigar. 


204  LABORATORY   GUIDE   IN 


Anaesthetics. 

The  most  important  members  of  the  group  of  drugs  used 
to  produce  general  anaesthesia  are.  'Chloroform  and  Ether. 
Their  depressant  action  on  the  central  nervous  system  has 
been  observed  in  some  of  the  experiments  previously  carried 
out  in  which  they  have  been  used  to  supplement  the  other 
drugs  used  as  anaesthetics  in  animals  (chloretone,  etc.).  The 
cardiac  effects  were  studied  in  the  frog,  rabbit  and  dog. 
In  these  animals  is  was  noticed  that  the  heart  dilated 
and  became  very  weak  before  there  was  very  much 
change  in  the  rate.  The  importance  of  this  in  practical 
anaesthesia  is  that  information  as  to  the  condition  of  the 
heart  cannot  be  derived  entirely  from  the  pulse  rate,  as  the 
heart  may  be  seriously  affected  and  yet  show  very  little 
change  in  the  nu'mber  of  its  contractions  per  minute.  The 
quality  of  the  pulse  must  also  be  noted. 

The  fact  that  chloroform  acts  very  much  more  strongly 
on  the  heart  than  ether,  was  also  demonstrated  in  the  ex- 
periments mentioned  above. 

Some  of  the  early  effects  of  the  anaesthetics  are  shown  in 
the  following  experiment : — 

I.  Place  a  rabbit  on  the  table  and  hold  it  by  placing  your 
hand  under  its  chest  in  such  a  way  that  you  can  count  its 
heart  and  respiration.  Pour  some  chloroform  on  a  little  ab- 
sorbent cotton  and  hold  it  to  the  animal's  nose  for  a  few 
moments  and  observe  any  change  in  the  rhythm  of  the  heart 
or  respiration. 

Repeat  the  experiment  using  ether  and  afterwards  am- 
monia. It  will  be  seen  that  the  same  effects  are  produced  by 


EXPERIMENTAL  PHARMACOLOGY..  205 


v^^l 


206  LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  207 

all  three  drugs.  They  are  caused  by  a  reflex  stimulation  of 
the  vagus  and  respiratory  centers  from  the  action  of  the  irri- 
tant fumes  on  the  mucous  membranes. 

Other   drugs  that  are  used  as  general   anaesthetics  are 
Nitrous  oxide  gas  and  Ethyl  chloride. 
14-p 


208  LABORATORY   GUIDE   IN 


Pilocarpus. 

•  r        «•»* 

Pilocarpine  is  an  alkaloid  derived  from  the  leaves  of  sev- 
eral species  of  Pilocarpus.  -It  has  .been  employed  in  certain 
of  the  earlier  experiments,  which  may  be  summarized  in 
order  to  give  a  more  complete  picture  of  the  action  of  the 
drug. 

Describe  the  effects  of  pilocarpine  on 

the  frog's  heart  (Exp.  page  135)  ; 
the  turtle's  heart  (Exp.  page.  144)  ; 
the  rabbit's  heart  (Exp.  page  136)  ; 
the  pupil  of  the  eye  (Exp.  page  188)  ; 
the  salivary  secretion  (Exp.  page  196)  ; 
and  on  the  intestinal  peristalsis  (Exp.  page  220). 
How  does  atropine  affect  the  activity  of  the  drug? 
I.     (These  drugs  will  be  furnished  you.)     Take  0.003  G. 
of  pilocarpine   hydrochloride   and   describe   any   symptoms 
noted.    Should  any  of  these  become  unpleasant  take  atropine 
sulphate  0.0005  G.  and  describe  any  changes. 

Other  drugs  which  resemble  pilocarpine  in  many  of  their 
actions  are  Muscarine,  from  one  of  the  poisonous  mush- 
rooms (Agaricus  muscarius),  and  Physostigmine  or  Eserine, 
from  the  Calabar  or  Ordeal  bean  (Physostigma  venenosum). 
Examine  the  leaves  of  Pilocarpus  jaborandi,  and  the 
calabar  bean. 

Therapeutic  Uses. 

The  employment  of  pilocarpine  in  diseases  of  the  eye  is 
discussed  elsewhere  (page  191).  The  drug  is  practically 
never  used  for  its  cardiac  action.  . 


EXPERIMENTAL  PHARMACOLOGY.  209 


(/^£^(2#^^ 


^    sfa^f^ 


^^^^       <^-L^^ 


x^s  ^ 


210  LABORATORY  GUIDE  IN 


BXP&RIMMNTAL  PHARMACOLOGY.  211 

It  is  used  to  increase  the  activity  of  the  sweat  glands 
(diaphoretic  action)  in  various  diseases  in  which  there  is 
an  abnormal  collection  of  fluid  in  the  body.  (Compare  Di- 
uretics, page  175).  Through  its  action  a  very  large  amount 
of  fluid  may  be  excreted  so  that  in  some  cases  the  body  may 
lose  from  four  to  nine  pounds  in  weight  from  a  single  dose 
of  the  drug. 

Pilocarpine  is  employed  in  some  diseases  of  the  kidneys  in 
which  the  urine  excretion  is  lessened,  with  the  idea  of  se- 
curing the  elimination  of  toxic  materials  by  the  sweat  glands. 


212  LABORATORY   GUIDE   IN 


Suprarenal  Gland  Extract. 

The  suprarenal  glands  contain  a  very  active  substance, 
Epinephrine,  which  can  be  isolated,  but  with  difficulty,  as  it  is 
very  unstable  and  easily  decomposed.  Various  preparations 
of  the  gland  are  put  on  the  market,  of  which  Adrenalin 
(Takarnine)  is  one  of  the  best  known. 

For  experimental  work  it  may  be  used  in  two  forms,  viz.:  A  solution  may 
be  prepared  by  boiling  a  minute  amount  of  the  powder  (adrenalin)  in  physi- 
ological salt  solution  until  the  fluid  becomes  a  light  pink.  The  objection  to  this 
method  is  that  it  does  not  allow  of  ex*act  dosage.  By  using  the  commercial 
adrenalin  chloride  (i-iooo)  this  objection  is  overcome.  From  one  to  three 
drops  of  this  preparation  give  good  results  in  experimental  work. 

Describe  the  effects  of  extract  of  the  suprarenal  gland  upon 

the  rabbit's  heart  (Exp.  page  139)  ; 

the  blood  pressure  (Exp.  page  151)  ; 

the  dog's  heart  (Exp.  page  159)  ; 

the  salivary  secretion  (Exp.  page  196)  ; 

the  kidney  vessels   (Exp.  page  168)  ; 

and  upon  intestinal  vessels  (Exp.  page  227). 
I.  Examine  the  local  effects  of  the  drug  by  putting  one 
drop  of  adrenalin  chloride  (i-iooo)  in  the  eye.  Compare 
with  the  normal  eye.  How  would  this  experiment  aid  you 
in  explaining  the  results  obtained  in  the  experiment  on  blood 
pressure  ? 

Therapeutic  Uses. 

The  powerful  effects  of  the  drug  on  the  circulation  are 
limited  in  their  application  to  therapeutics  by  their  transitory 
nature  and  by  the  fact  that  in  order  to  act  on  the  heart  or 
blood  vessels  (except  the  local  action),  the  drug  has  to  be 


EXPERIMENTAL  PHARMACOLOGY.  213 


214  LABORATORY  GUIDE  IN 


EXPERIMENTAL   PHARMACOLOGY.  215 

injected  intravenously.  In  some  cases  of  shock  and  collapse 
it  has  been  given  in  this  way,  but  it  requires  constant  admin- 
istration as  the  effects  pass-  off  so  rapidly.  It  is  of  most 
value  for  local  application  in  surgical  operations  to  constrict 
the  vessels  and  thus  render  the  field  of  operation  bloodless. 


216  LABORATORY   GUIDE,   IN 


The  Nitrite  Series. 

This  series  comprises  several  drugs  whose  action  in  the 
body  is  confined  almost  exclusively  to  the  blood  vessels.  The 
principal  members  are  Amyl  Nitrite,  Nitro  glycerin,  Sodium 
and  Potassium  Nitrite. 

They  have  been  employed  in  various  experiments  and  the 
results  are  to  be  collected  and  summarized. 
Describe  the  effects  of  amyl  nitrite  on — 

the  rabbit's  blood  pressure  (Exp.  page  151)  ; 
the  vessels  of  the  kidney  (Exp.  page  232)  ; 
and  on  the  vessels  of  the  intestine  (Exp.  page 

227). 

Describe  the  effects  on  the  human  blood  pressure  of 
amyl  nitrite  (Exp.  page  200)  ; 
nitroglycerin  (Exp.  page  203)  ; 
sodium  nitrite  (Exp.  page  203), 
and    of   sodium   nitrite   on   the   excised   kidney 
vessels  (Exp.  page  167). 

Therapeutic  Uses. 

The  members  of  this  series  are  employed  in  medicine  to 
lower  the  blood  pressure  and  to  relieve  vascular  spasm. 


EXPERIMENTAL  PHARMACOLOGY.  217 


218  LABORATORY   GUIDE  IN 


EXPERIMENTAL   PHARMACOLOGY.  219 


The  following  experiments  are  not  suitable  for  the  regu- 
lar laboratory  course,  but  may  be  carried  out  by  advanced 
students ;  they  will  also  serve  very  well  for  class  demonstra- 
tions by  the  instructor  in  charge. 

Intestinal  Peristalsis. 

The  peristaltic  movements  of  the  intestines  and  the  effects 
of  various  drugs  upon  them  may  be  demonstrated  very  well 
by  submerging  an  animal  in  a  saline  solution,  opening  the 
abdomen  and  keeping  the  intestines  under  the  surface  of  the 
water  by  means  of  a  pane  of  glass. 

For  this  experiment  a  metallic  tank  is  needed  which  will 
be  large  enough  to  allow  a  rabbit  to  be  placed  in  it  when 
the  animal  is  tied  to  an  operating  board.  A  tank  measuring 
70x30x30  cm.  is  large  enough  for  most  purposes  and  it 
should  be  mounted  on  legs  long  enough  to  allow  a  Bunsen 
burner  to  be  placed  under  it.  The  operating  board  should 
be  weighted  with  lead  so  that  it  will  sink  in  the  water.  A 
pane  of  glass  to  cover  the  intestines  and  some  small  lead 
piping,  which  can  be  bent  over  the  sides  of  the  tank  to  act 
as  a  support  for  the  glass,  and  a  bath  thermometer  complete 
the  special  apparatus  necessary  for  the  experiment. 

Fill  the  tank  about  two-thirds  full  of  water  and  add 
sodium  chloride  to  make  a  physiological  salt  solution  (8  G. 
per  liter) .  Light  the  Bunsen  burner  under  the  tank  and  heat 
the  solution  until  it  is  of  the  body  temperature. 


220  LABORATORY   GUIDE   IN 

Operation. 

In  the  meantime  anaesthetize  a  rabbit  in  the  usual  manner 
and  when  anaesthesia  is  complete  insert  a  venous  cannula 
in  the  jugular  vein  and  also  insert  a  tracheal  cannula.  Place 
short  pieces  of  rubber  tubing  on  both  free  ends  of  the 
tracheal  cannula  and  close  one  by  means  of  a  clamp.  The 
second  piece  of  tubing  is  to  extend  above  the  surface  of  the 
water  when  the  animal  is  submerged. 

Place  the  animal  in  the  warm  salt  solution  which  should 
completely  cover  it.  Take  great  care  no  water  enters  the 
tracheal  tube,  the  open  end  of  which  is  to  be  tied  to  a 
support  above  the  tank.  Make  an  incision  through  the  skin 
and  muscles  in  the  median  line  of  the  abdomen  from  the  end 
of  the  sternum  to  the  symphysis.  The  abdominal  walls  may 
be  drawn  outward  and  secured  to  the  operating  board  by 
means  of  tacks.  The  intestines  will  float  to  the  top  of  the 
water  and  they  must  be  held  just  under  the  surface  by 
placing  the  pane  of  glass  over  them  and  supporting  the  latter 
in  position  by  means  of  the  lead  tubing. 

The  peristaltic  movements  may  now  be  observed  as  well 
as  the  vascular  changes  which  will  follow  the  injection  of 
the  drugs  named  below ;  the  changes  in  color  of  the  intestines 
indicate  the  alterations  in  the  circulation  of  the  blood. 

Inject  into  the  jugular  vein  the  following  drugs  in  the 
order  named : — 

Nicotine  chloride,  5  mg., 

Pilocarpine  hydrochloride,  5  mg., 

Atropine  sulphate,  I  mg., 

Veratrine  sulphate,  i  mg., 

Adrenalin  chloride  (i-iooo  sol.),  2  drops, 

Amyl  nitrite,  2  or  3  drops  given  by  inhalation. 


EXPERIMENTAL  PHARMACOLOGY.  221 


222  LABORATORY   GUIDE    IN 


EXPERIMENTAL   PHARMACOLOGY.  223 


Vasomotor  Changes  in  the  Intestine. 

The  special  apparatus  needed  for  this  experiment  is,  first, 
some  form  of  intestinal  plethysmograph  and  second,  an  in- 
strument to  record  the  intestinal  vascular  changes  upon  a 
blackened  drum. 

The  plethysmograph  may  be  made  very  easily  of  plaster 
of  Paris  after  the  pattern  devised  by  A.  Edmunds.1 

With  the  plaster  make  a  hemispherical  basin  ten  centi- 
meters in  diameter  and  four  deep  with  the  walls  about  seven 
millimeters  thick  and  with  an  elliptical  opening  in  the  bottom 
of  the  basin  three  centimeters  by  one  and  a  half.  Make  a 
small  opening  in  one  side  of  the  apparatus  and  cement  in  it 
a  short  piece  of  glass  tubing  which  connects  with  the  interior 
of  the  basin  and  which  extends  outside  three  or  four  centi- 
meters to  allow  of  rubber  pressure  tubing  being  connected 
with  it.  Smooth  off  the  upper  edge  of  the  apparatus  with 
sand  paper  so  that  a  piece  of  glass  will  fit  fairly  tightly  on  it. 
A  plethysmograph  made  in  this  way  will  answer  all  the  pur- 
poses of  the  experiment.  Its  more  complicated  forms  may 
be  made  as  described  in  the  article  referred  to.  A  glass  plate 
which  will  close  the  top  of  the  plethysmograph  is  needed  as 
well  as  a  piece  of  pressure  tubing  long  enough  to  connect  the 
plethysmograph  with  the  recording  apparatus,  which  may  be 
either  a  tambour,  a  delicate  piston  recorder  or  a  Brodie 
bellows  recorder.  Either  of  these  should  be  provided  with  a 
side  tube  to  allow  of  proper  equalization  of  pressure  on  both 
sides  of  the  piston  or  membrane  of  the  recorder. 

Vaseline  is  needed  as  well  as  a  petrolate  which  melts  at 


r.  of  Physiol.,  Vol.  XXII,  1898,  page  380. 

15-p 


224  LABORATORY   GUIDE  IN 

a  higher  temperature  than  vaseline  and  which  may  be  made 
by  melting  the  latter  with  wax,  so  that  the  resulting  mixture 
is  fairly  solid  when  cold. 

Operation. 

Anaesthetize  a  rabbit  or  a  cat  as  usual,  tie  it  on  an  operat- 
ing board  and  insert  a  venous  cannula. 

If  it  is  desired  to  take  a  tracing  of  the  carotid  blood  pres- 
sure at  the  same  time,  the  artery  must  be  isolated,  a  cannula 
inserted  and  connected  with  the  mercury  manometer  accord- 
ing to  the  directions  given  on  page  148.  By  thus  taking 
simultaneous  tracings  of  the  general  blood  pressure  and  of 
the  vascular  conditions  in  the  intestinal  area  very  instructive 
records  may  be  obtained. 

It  is  best  to  warm  the  plethysmograph  slightly  before  ap- 
plying it  and  to  have  plenty  of  hot  salt  solution  ready  for  use 
as  well  as  some  absorbent  cotton.  Make  an  incision  about 
4  cm.  long  in  the  median  line  of  the  abdomen  about  midway 
between  the  sternum  and  symphysis  and  with  the  fingers 
carefully  pull  out  of  the  abdominal  cavity  a  short  loop  of  the 
small  intestine.  Place  the  elliptical  opening  of  the  plethys- 
mograph over  the  incision  and  intestinal  loop  and  support  the 
instrument  in  place  with  an  iron  ring.  Draw  the  loop  of  the 
intestine  into  the  apparatus  until  a  section  from  20  to  30  cm. 
long  is  lying  in  the  basin.  Cover  the  intestine  temporarily 
with  absorbent  cotton  dipped  in  the  hot  salt  solution.  Ligate 
doubly  each  end  of  the  section  of  intestine  and  cut  across  it 
between  the  ligatures,  tearing  down  the  mesentery  as  far  as 
possible  and  ligating  any  bleeding  vessels.  Now  return  to  the 
abdominal  cavity  the  upper  and  lower  attached-  ends  of  the 
intestine,  leaving  the  isolated  loop  in  the  plethysmograph 


EXPERIMENTAL  PHARMACOLOGY.  225 


226  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  227 

and  connected  to  the  animal  by  its  section  of  mesentery  con- 
taining the  vessels  and  nerves. 

Pack  absorbent  cotton  rubbed  up  with  the  hard  vaseline 
around  the  mesentery  so  as  to  completely  close  the  elliptical 
opening,  using  the  soft  vaseline  to  make  it  air  tight  near  the 
vessels ;  great  care  must  be  taken  not  to  constrict  the  vessels. 
Remove  the  cotton  covering  from  the  intestines ;  vaseline  the 
upper  rim  of  the  plethysmograph  and  close  it  in  with  the 
pane  of  glass  which  has  been  dipped  in  warm  salt  solution  to 
prevent  the  condensation  of  moisture  on  its  surface. 

Connect  the  instrument  with  the  recorder,  close  the  side 
tube  on  the  latter  and  take  a  tracing,  which,  if  the  experi- 
ment has  been  properly  carried  out,  will  show  both  cardiac 
beats  and  the  respiratory  waves.  Arrange  the  lever  to  write 
in  line  with  the  blood  pressure  pointer  and  the  time  marker 
which  has  been  placed  below. 

Take  tracings  to  show  the  effects  of  the  following  drugs 
on  the  general  arterial  blood  pressure  and  on  the  vessels  of 
the  splanchnic  area. 

Adrenalin  chloride  (i-iooo),  3  drops  in  salt  solution. 

Digitalis,  tincture,  I  cc.  in  salt  solution. 

Nicotine  chloride,  5  mg.  in  salt  solution. 

Amyl  nitrite,  2  drops  to  be  inhaled,  on  cotton. 


228  LABORATORY   GUIDE   IN 


Vascular  Changes  in  the  Kidney. 

To  record  the  changes  taking  place  in  the  size  of  the  kid- 
ney due  to  the  differences  in  its  blood  content,  an  oncometer 
and  a  recording  apparatus  are  needed.  The  former  can  be 
made  from  plaster  of  Paris  in  the  same  manner  as  the  intes- 
tinal plethysmograph.  Several  sizes  should  be  made  to  ac- 
commodate the  kidneys  of  different  sized  animals.  A  cylin- 
drical-shaped oncometer  with  a  flat  base,  having  a  diameter 
of  about  5  cm.  and  with  walls  about  3.5  cm.  high  is  large 
enough  for  the  average  sized  cat  or  rabbit.  One  side  should 
have  a  cut  in  it  about  0.5  cm.  wide  and  extending  nearly  to 
the  bottom  of  the  apparatus,  where  the  cut  should  be  widened 
to  allow  greater  room  for  packing  around  the  pedicle  of  the 
kidney.  On  the  opposite  side  of  the  oncometer  is  a  glass 
tube  communicating  with  the  interior  of  the  apparatus  and 
extending  outward  for  the  attachment  of  rubber  pressure 
tubing.  A  piece  of  glass  is  needed  to  fit  tightly  the  top  of 
the  instrument.  Vaseline  together  with  a  mixture  of  vaseline 
and  wax  (as  described  on  page  223)  is  also  necessary. 

For  recording  the  volume  changes  on  the  kymograph,  a 
tambour,  piston  recorder,  or  a  bellows  recorder  as  mentioned 
under  the  experiment  on  the  intestinal  vessels  (page  223) 
is  required.  It  should  be  fitted  as  before  with  a  side  tube. 

Operation. 

Anaesthetize  a  rabbit  or  a  cat  and  insert  one  cannula  in  the 
jugular  vein  and  a  second  in  the  carotid  artery.  Connect  the 
latter  with  the  mercury  manometer  for  blood  pressure, 
which  has  been  arranged  according  to  the  directions  given 


EXPERIMENTAL  PHARMACOLOGY.  229 


230  LABORATORY   GUIDE   IN 


EXPERIMENTAL  PHARMACOLOGY.  231 

on  page  148.  Either  kidney  may  be  employed  in  the  experi- 
ment, the  choice  largely  depending  upon  the  arrangement  of 
the  apparatus  and  the  light.  If  the  left  has  been  selected, 
make  an  incision  in  the  abdomen  well  over  in  the  left  flank, 
extending  from  the  costal  margin  downward,  the  length  of 
the  incision  necessary  depending  upon  the  size  of  the  onco- 
meter.  Draw  the  side  muscles  outward,  if  necessary  con- 
tinuing the  cut  in  the  muscles  along  the  costal  margin,  and 
nail  the  reflected  muscles  to  the  operating  board.  The  intes- 
tines and  stomach  should  be  packed  back  with  absorbent 
cotton  which  has  been  dipped  in  hot  salt  solution  and  the 
abdominal  cavity  closed  in  as  far  as  possible  with  a  hot 
towel.  This  should  leave  the  left  kidney  well  exposed. 
Loosen  it  very  carefully  from  all  its  fibrous  attachments  and 
remove  the  fat  as  far  as  possible,  so  that  the  kidney  remains 
connected  only  by  the  structures  entering  the  hilus. 

Place  the  organ  in  the  warmed  oncometer  so  that  the 
pedicle  passes  through  the  widened  end  of  the  cut  in  the 
side  of  the  apparatus.  Pack  around  the  pedicle  with  ab- 
sorbent cotton  saturated  with  the  hardened  vaseline  and 
completely  close  the  opening  in  the  side  in  the  same  way, 
employing  the  soft  vaseline  to  make  it  entirely  air  tight.  Of 
course  it  is  essential  to  see  that  no  undue  pressure  is  exerted 
on  the  kidney  vessels ;  the  circulation  must  remain  perfectly 
free  in  them.  Close  in  the  top  of  the  oncometer  with  the 
pane  of  glass  which  is  made  air  tight  with  vaseline.  Con- 
nect the  outlet  tube  of  the  instrument  with  pressure  tubing, 
the  other  end  of  which  is  connected  with  the  recorder.  Ar- 
range the  writing  point  of  the  recorder  so  that  it  marks 
directly  above  the  pointer  of  the  blood  pressure  manometer 
and  the  time  marker.  Close  the  side  tubes  of  the  recorder 


232  LABORATORY   GUIDE   IN 

and  the  lever  should  show  the  distinct  cardiac  pulsations  as 
they  are  indicated  by  the  changes  in  the  kidney  volume.  If 
the  pulsations  do  not  show,  either  the  kidney  vessels  are  con- 
stricted or  the  apparatus  is  not  air  tight. 

When  all  the  apparatus  is  working  satisfactorily  inject  the 
following  drugs  into  the  jugular  vein : 

Adrenalin  chloride,  i-iooo,  I  to  3  drops  in  salt  solution. 

Tincture  of  digitalis,  I  cc.  in  salt  solution. 

Nicotine  chloride,  5  mg.  in  salt  solution. 

Amyl  nitrite,  3  drops,  given  by  inhalation. 


EXPERIMENTAL   PHARMACOLOGY.  233 


234  LABORATORY   GUIDE  IN 


EXPERIMENTAL  PHARMACOLOGY.  235 


List  of  Drugs  and  Reagents  Required, 

Acacia,  powdered. 
Acids : 

Hydrochloric,  con. 

lodic,  5%. 

Nitric,  con. 

Phosphotungstic,  2%. 

Picric,  saturated. 

Sulphuric,  con. 

Sulphuric,  10%. 

Tannic,  10%. 
Aconite,  tincture. 
Adrenalin  ( Takamine ) . 
Adrenalin  chloride,  i-iooo. 
Almonds,  bitter. 
Alcohol,  95%. 
Ammonium  hydrate,  10%. 
Amygdalin. 
Amyl  nitrite. 
Asafoetida. 

Atropine  sulphate,  0.1%  solution. 
Atropine  sulphate,  for  hypoderm.  use. 
Belladonna,  No.  60  powder. 
Caffeine,  cryst. 
Caffeine,  2%  solution. 
Calcium  chloride. 
Charcoal. 
Chloral. 
Chloretone. 


236  LABORATORY   GUIDE   IN 

Chlorine  water. 

Chloroform. 

Cocaine  hydrochloride,  4%  sol. 

Curara,  standardized  sol.   (page  88). 

Digitalis  leaves,  No.  60  powder. 

Digitalis  leaves,  bruised. 

Ether. 

Fehling's  solution.    U.  S.  P. 

Ferric  chloride. 

Iodine  in  potassium  iodide.    U.  S.  P. 

Lycopodium. 

Manganese  dioxide. 

Mercury-potassium  iodide.    U.  S.  P. 

Milk  sugar. 

Morphine  sulphate. 

Nicotine  chloride.     (See  note,  page  92). 

Nux  vomica,  No.  60  powder. 

Oils: 

Castor. 

Cotton  seed. 

Turpentine. 
Opium. 
Paraldehyde. 

Pilocarpine  hydrochloride,  0.5%  sol. 
Pilocarpine  hydrochloride,  0.02  G.  tablets. 
Potassium  bichromate. 

Carbonate,  10%. 
chloride, 
hydrate,  10%. 

"  hydrate,  10%,  in  absol.  alcohol. 

Quinine. 


EXPERIMENTAL  PHARMACOLOGY.  237 


238  LABORATORY   GUIDE   IN 


EXPERIMENTAL   PHARMACOLOGY.  239 

Quinine  hydrochloride. 

"       sulphate. 
Sodium  chloride,  cryst. 
nitrate,  $%  sol. 
"       nitrite,  cryst. 
"       sulphate,  sat.  sol. 
Starch. 
Strychnine  sulphate,  0.1%  sol. 

sulphate,  0.002  G.  tablets. 
Sugar. 
Sulphonal. 

Veratrine  sulphate,  0.1%  sol. 
16-p 
I 


INDEX. 

PAGE. 

Absinthe  79 

Aconite   I2^ 

Aconitine  I2° 

Acrole'in    4° 

Adrenalin    2I2 

Alkaloids  24 

Alkaloidal  precipitants   3* 

Amygdalin   32 

Amyl   nitrite    216 

Anaesthetics 204 

Anaesthetic  for  cat 12 

for  dog 12 

for  rabbit 1 1 

Analysis  of  tracings 160 

Apalache  tea 96 

Aqua  4° 

Assay  of  tincture  of  digitalis 120 

of  the  infusion  of  digitalis 124 

Atropine    179 

isolation  of 180 

Balsams   43 

Belladonna  series 179 

Berberine 32 

Bladder  cannula,  to  insert, 171 

Blood  pressure  in  man,  to  estimate 199 

drugs  on  200 

Blood  pressure  in  animals,  drugs  on 147 

Brncine  64 

Caffeine       96 

Camphor    79 

Cat  box 12 

Cannulas,  arterial   20 

tracheal   23 

venous   19 

241 


242  INDEX. 

PAGE. 

Chemistry  of  drugs 24 

Chloral   51 

Chloralose  55 

Chloretone   .  .- 55 

Chloroform  204 

Chorda  tympani  nerve,  to  find 191 

Cinchona 103 

Clonic  convulsion  79 

Cocaine  100 

Cocaine  on  pupil 188 

Codeine  80 

Coffee   96 

Coniine  32 

Continental  method  of  emulsification HI 

Curara  88 

Curine   88 

Deadly  nightshade  179 

Diaphoretic    208 

Digitalin   36 

Digitalis  119 

Digitalis  on  kidney 172 

Digitonin  124 

Digitoxin    36 

Diuresis'  168 

Diuretics    175 

Diuretic,  cardiac 128 

Dog's  heart,  drugs  on 151 

Dover's  powder 87 

Drugs  and  reagents  required 235 

Emulsifiers 108 

Emulsin 35 

Emulsions   108 

Epinephrine 212 

Eserine 208 

Eserine  on  pupil 188 

Essential  oils 36 

Ether    204 

Ethereal   oils 36 

Ethyl  chloride 207 

Excipient  115 

Extract   179 


INDEX.  243 

PAGE. 

Fehling's  reaction  31 

Fixed  oil  emulsion 1 1 1 

Fluid  extract   64 

Frog,  operations  on.     See  Operations. 

Frog's  heart,  drugs  on 132 

Glucosides    32 

Guarana  paste  96 

Gums  43 

Gum  resins  43 

Gum  resin  emulsion m 

Henbane    179 

Homatropine , 191 

Hyoscine   179 

Hyoscyamine    179 

Hypnotics  51 

Infusion  of  digitalis . . 124 

Isolation  of  strychnine 67 

of  atropine 180 

Intestinal  peristalsis 219 

Kola  nut   96 

Laudanum    87 

Mammals,  operations  on.     See  Operations. 

Meconic  acid   80 

Meconic  acid  reaction 84 

Monk's  hood  128 

Morphine 80 

Murexoin  test   99 

Muscarine   208 

Mydriasis    179 

Mydriatics    179, 191 

Myocardiograph  for  turtle 144 

for  dog 155 

Myotics  191 

Nicotine    92 

Nicotine  chloride    footnote  92 

Nicotine  on  sympathetic  nerves 187 


244  INDEX. 

PAGE. 

Nitrite  series  216 

Nitrite,  amyl    216 

sodium   216 

potassium   216 

Nitroglycerin    216 

Nitrous  oxide  207 

''Non-official"    drugs    7 

Nux   vomica    64 

''Official"  drugs  7 

Oils,  fatty   36 

fixed    36 

volatile    36 

Oil  of  bitter  almonds 35 

Oleoresins    43 

Oncometer  for  kidney 228 

Opium  80 

Operations  on  frogs,  pithing  15 

to  inj  ect  in  lymph  sac  15 

to  expose  the  heart 16 

Operations  on  mammals,  to  insert  an  arterial  cannula 20 

to  insert  a  tracheal  cannula 23 

to  insert  a  venous  cannula 19 

Paraguay  tea 96 

Paraldehyde       51 

Paregoric 87 

Percolation   .-  59 

Perfusion  of  kidney 167 

Pharmacopoeia    7 

Physiology  of  frog's  nervous  system  44 

Physostigmine   208,  188 

Pills   112 

Pill  mass 115 

Pilocarpine  on  pupil 188 

Pilocarpus 208 

Pipette,  glass  injecting 15 

Pithing  frogs 15 

Plethysmograph,  intestinal 223 

Quinine    103,  24 

Quinine  sulphate 24 


INDEX.  245 

PAGE. 

Rabbit's  heart,  drugs  on 135 

Reagents  and  drugs  required 235 

Resins    43 

Sage  79 

Salicin    36 

Saline  diuretic ".  175 

Salivary  secretion 191 

Salivary  secretion,  drugs  on 195 

Seed  emulsion 108 

Shaking-out  process 31 

Soporifics    51 

Sodium  chloride,  on  kidney 175 

Sodium  nitrate,  on  kidney 172 

Sodium  nitrite,  on  kidney  vessels 168 

Sphygmomanometer    199 

Spiritus  40 

Squills 119 

Stomach  tube  for  cats 1 1 

dogs   12 

rabbits   1 1 

Strophanthus 119 

Strychnine    64 

Submaxilliary  duct,  to  find 191 

Sulphonal  51 

Suprarenal  gland  extract 212 

Sympathetic  system,  drugs  on 187 

Syrupus    40 

Takamine   212 

Tansy   79 

Tea 96 

Tetronal 55 

Theobromine  96 

Theophyllin    96 

Thornapple   179 

Thebaine 87 

Thujon  79 

Tincture • •  •  IJ9 

Tobacco  92 

Tonic  79 

Tonic  convulsion  71 

Trional  .                                                     55 


246  INDEX. 

PAGE. 

Turtle's  heart,  drugs  on 139 

Tubocurarine    88 

Urethane 55 

Vasomotor  effects  in  the  intestine  223 

in  the  kidney 228 

Vagus  nerve,  in  frog 132 

in  turtle  143 

Veratrine  95,  131 

Vitali's  test 183 

Volatile  oils  36 


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at  the  University  of  Michigan,  President  of  the  University  of  Washington, 
and  Wm.  J.  Hussey,  Assistant  Professor  of  Astronomy  in  the  Leland 
Stanford,  Jr.  University  8vo.  294  pages.  $3.50. 

GREENE* — The  Action  of  Materials  Under  Stress,  or  Structural  Mechanics. 
With  examples  and  problems.  By  Charles  E.  Greene,  A.  M.,  M.  E., 
Professor  of  Civil  Engineering  in  the  University  of  Michigan.  Consulting 
Engineer.  Octavo.  Cloth,  $3.00. 

GUTHE. — Laboratory  Exercises  "with  Primary  and  Storage  Cells.  By  Karl  E. 
Guthe,  Assistant  Professor  of  Physics,  University  of  Michigan.  58  pages. 
1 7  illustrations.  Cloth,  45  cents. 

HERDMAN-NAGLER. — A  Laboratory  Manual  of  Electrotherapeutics.  By 
William  James  Herdman,  Ph.  B.,  M.  D.,  Professor  of  Diseases  of  the 
Nervous  System  and  Electrotherapeutics,  University  of  Michigan,  and  Frank 
W.  Nagler,  B.  S.,  Instructor  in  Electrotherapeutics,  University  of  Michigan. 
Octavo.  163  pages.  55  illustrations.  Cloth,  $1.50. 

HUBER* — Directions  for  Work  in  the  Histological  Laboratory.  By  G.  Carl  Huber, 
M.  D.,  Assistant  Professor  of  Histology  and  Embryology,  University  of 
Michigan.  Third  edition,  revised  and  enlarged.  Octavo.  204  pages. 
Cloth,  $1.50. 

MONTGOMERY  -  SMITH, — Laboratory  Manual  of  Elementary  Chemistry.  By 
Jabez  Montgomery,  Ph.  D.,  Professor  of  Natural  Science,  Ann  Arbor  High 
School,  and  Roy  B.  Smith,  M.  S.,  Colgate  University.  12  mo.  400  pages. 
Cloth,  #1.25. 

MORRIS. —  Clinical  Temperature  Chart.  By  Roger  S.  Morris,  A.  B.,  M.  D., 
University  of  Michigan.  Postpaid,  48  cents  per  quire. 

There  are  spaces  for  stools,  urine,  dates  and  days  of  disease,  besides  abundant  room  for 
the  personal  data,  diagnosis,  treatment,  termination,  and  for  daily  notes.  Anything  that 
assists  and  favors  accuracy  in  clinical  records,  either  in  hospital  or  private  practice,  should 
be  welcomed  by  physicians,  and  those  who  are  looking  for  a  chart  different  from  those  now 
in  use  are  warmly  advised  to  examine  this  one. 

NETTO* — The  Theory  of  Substitutions  and  its  Application  to  Algebra.  By  Dr. 
Eugene  Netto,  Professor  of  Mathematics  in  the  University  of  Giessen. 
Revised  by  the  author  and  translated  with  his  permission,  by  F.  N.  Cole, 
Ph.  D.,  formerly  Assistant  Professor  of  Mathematics  in  the  University  of 
Michigan,  Professor  of  Mathematics,  Columbia  University.  8  vo.  301 
pages.  Cloth,  $3.00. 

NOVY. — Laboratory  Work  in  Physiological  Chemistry.  By  Frederick  G.  Novy, 
Sc.  D.,  M.  D.,  Junior  Professor  of  Hygiene  and  Physiological  Chemistry, 
University  of  Michigan.  New  third  edition  preparing. 

NOVY. — Laboratory  Work  in  Bacteriology.  By  Frederick  G.  Novy,  Sc.  D., 
M.  D.,  Junior  Professor  of  Hygiene  and  Physiological  Chemistry,  University 
of  Michigan.  New  third  edition  preparing. 

REED-GUTHE. — A  Manual  of  Physical  Measurements.  By  John  O.  Reed, 
Junior  Professor  of  Physics,  University  of  Michigan,  and  Karl  E.  Guthe, 
Assistant  Professor  of  Physics,  University  of  Michigan.  185  pages,  89  illus- 
trations. Octavo.  $1.50. 


SOLIS. —  The  Diagnosis  of  Disease*  of  the  Cord,  Location  of  Lesions.  By  Dr.  Grasset. 
Translated  by  Jeanne  C.  Soils,  M.  D.,  Demonstrator  of  Nervous  Diseases  and 
Electrothereapeutics  in  the  University  of  Michigan.  98  pages.  Cloth, 
65  cents. 

VAUGHAN. — Contributions  to  Medical  Research,  Dedicated  to  Victor  Clarence 
Vaughan.  By  Colleagues  and  Former  Students  of  the  Department  of  Med- 
icine and  Surgery  of  the  University  of  Michigan,  on  the  Twenty-fifth  Anni- 
versary of  his  Doctorate.  Large  octavo  size.  640  pages  and  200  illustrations. 
Bound  in  full  Cloth,  $5.00;  Half  Morocco,  $6.00. 

WARTHIN. — Practical  Pathology  for  Students  and  Physicians.  A  Manual  of 
Laboratory  and  Post-Mortem  Technic,  Designed  Especially  for  the  Use  of 
"Junior  and  Senior  Students  in  Pathology  at  the  University  of  Michigan.  By 
Aldred  Scott  Warthin,  Ph.D.,  M.  D.,  Instructor  in  Pathology,  University  of 
Michigan.  Octavo.  234  pages.  Cloth,  $1.50. 

WATSON. —  Tables  for  the  Calculation  of  Simple  or  Compound  Interest  and  Discount 
and  the  Averaging  of  Accounts.  The  Values  of  Annuities,  Leases,  Interest  n 
Estates  and  the  Accumulations  and  Values  of  Invstments  at  Simple  or  Com- 
pound Interest  for  all  Rates  and  Periods^  also  Tables  for  the  Conversion  of 
Securities  and  Values  of  Stocks  and  Bonds.  With  full  explanation  for  Use. 
By  James  C.  Watson, 'Ph.D.,  LL.D.  Quarto.  Cloth,  $2.50. 

WRENTMORE.— Plain  Alphabets  for  Office  and  School.  Selected  bv  C.  G. 
Wrentmore,  B.S.,  C.E.,  Instructor  in  Descriptive  Geometry  and  Drawing, 
University  of  Michigan.  Oblong.  19  plates.  Half  leather,  75  cents. 

WRENTMORE-GOULDINO.— A  Text  Book  of  Elementary  Mechanical  Draw- 
ing for  Use  in  Office  or  School.  By  Clarence  G.  Wrentmore,  B.S.  C.E., 
and  Herbert  J.  Goulding,  B.S.,  M.E.,  Instructors  in  Descriptive  Geometry 
and  Drawing  at  the  University  of  Michigan.  Quarto.  109  pages  and  165 
cuts.  $1.00. 

Souvenir  of  the  University  of  Michigan,  Ann  Arbor.  Containing  38  photo- 
gravures. President  James  B.  Angell,  prominent  University  Buildings, 
Fraternity  Houses,  Churches,  Views  of  Ann  Arbor,  Etc.,  Etc.  Bound  in 
blue  vellum  and  brown  ooze  leather.  Price,  50  cents  and  $1.00,  postpaid. 

Physical  Laboratory  Note  Book. — A  Note  Book  for  the  Physical  Laboratory. 
Designed  to  be  used  in  connection  with  any  Physical  Laboratory  Manual. 
Contains  full  directions  for  keeping  a  Physical  Laboratory  Note  Book.  112 
pages  of  excellent  ledger  writing  paper,  ruled  in  cross  sections,  Metric  System, 
size  7x9^  inches.  Bound  in  full  canvas,  leather  corners.  Price,  by  mail, 
30  cents.  Special  prices  to  Schools  furnished  on  application. 

Botanical  Laboratory  Note  Book. — A  Note  Book  for  the  Botanical  Laboratory. 
200  pages  of  best  writing  paper,  ruled  with  top  margins.  Pocket  on  inside  of 
front  cover  for  drawing  cards.  Bound  in  substantial  cloth  cover  and  leather 
back.  Size  6x9^.  Price,  by  mail,  35  cents.  Special  prices  to  Schools 
furnished  on  application. 

Engineering  Laboratory  Note  Book. — A  Note  Book  for  the  Engineering  Labor- 
atory. University  of  Michigan.  Full  sheep  binding.  Size  5^x8.  Con- 
tains 200  pages.  ( With  general  directions.  Cross  section  ruled.  )  Price, 
75  cents. 

Field  Engineering  Note  Book,  Surveying. — 200  pages.  Cross  section  ruled. 
Full  sheep  binding,  Co  cents. 


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DATE  DUE  SLIP 

UNIVERSITY  OF  CALIFORNIA  MEDICAL  SCHOOL  LIBRARY 

THIS  BOOK  IS  DUE  ON  THE  LAST  DATE 
STAMPED  BELOW 


SEP  1  4  1950 


2m-5,'31 


.(Y 


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